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Effect of insulin on glucose and amino acid uptake by skeletal muscle following burn injury. Studies with 2-deoxyglucose and alpha-aminoisobutyric acid.

Basal and insulin-stimulated uptake of 2-deoxyglucose (2-DG) and alpha-aminoisobutyric acid (AIB) by cells of the intact soleus muscle were studied 3 days after a 3-second scald on one hind limb of the rat. Soleus muscles from the burned and unburned limbs of burned rats, as well as from controls, were incubated with 1 mM [14C]2-DG or 50 micro M [14C] AIB and insulin (0, 1, 1.0, 10.0, or 100.0 mU/ml). Extracellular space was measured with [3H]inulin. Basal uptake of 2-DG and AIB by the cells of the burned limb muscles were 68% (p less than 0.001) and 40% (p less than 0.05), respectively, higher than those of controls. The intracellular concentrations of 2-DG and AIB of burned limb muscles were 112% (p less than 0.01) and 72% (p less than 0.01), respectively, higher than those in control muscles. Corresponding values in soleus muscles from the unburned limb of burned rats did not differ from controls. Addition of insulin increased both the rates of cellular uptake and intracellular concentrations of 2-DG and AIB in control and burned limb muscles but did not significantly affect those of burned limb muscles. It is concluded that thermal injury suppresses the insulin-induced augmentation of 2-DG and AIB uptake by skeletal muscle underlying the burn wound but does not alter the insulin sensitivity of skeletal muscle in the unburned region. Since glucose and amino acid uptakes are stimulated by insulin independently, the cellular mechanism of the burn-induced unresponsiveness to insulin may be located at a site which is common to both processes.

Amino Acids↗

Urinary pseudouridine and beta-aminoisobutyric acid in patients with low grade urothelial tumours. Relations between excretion and tumour recurrence.

The simultaneous 24 hours excretion of pseudouridine and beta-aminoisobutyric acid in the urine from patients treated for low grade urothelial tumours has been determined and related to tumour recurrence inside of 6 months after the determinations. The results of 53 assays in 39 patients without clinical signs of recurrence at the time of the detrmination showed a high excretion of pseudouridine in 53% and of beta-aminoisobutyric acid in 28.5% of the assays. Recurrences appeared more often after a high urinary pseudouridine (53.5%) than after a low, but the difference was not statistically significant (p more than 0.05) and more often after a low urinary beta-aminoisobutyric acid (52.5%) than after a high (p less than 0.01). The highest incidence of recurrence was in patients with a simultaneously high urinary pseudouridine and a low urinary beta-aminoisobutyric acid. Seventy per cent of these excretion patterns were from patients, who developed a recurrence before 6 months (p less than 0.002).

Adult↗

The stereochemistry of peptides containing alpha-aminoisobutyric acid.

The introduction of alpha-aminoisobutyric acid (Aib) into peptides dramatically limits the range of accessible backbone conformations. The presence of two geminal methyl groups at C alpha sterically compels Aib residues to largely favor structures in the right- or left-handed 3(10)/alpha-helical regions (phi approximately +/- 60 +/- 20 degrees, psi approximately +/- 30 +/- 20 degrees) of the peptide conformational map. Aib residues occur extensively in microbial peptides which form transmembrane channels. This observation has stimulated considerable interest in the stereochemistry of Aib peptides. This review summarizes theoretical studies on the conformations of Aib residues and examines the available data on solid-state structures, derived from single crystal X-ray diffraction studies. Crystal structures of over three dozen Aib-containing peptides, ranging in length from 2 to 11 residues, have been reported so far which exemplify various types of beta-turns, consecutive beta-turns, and helical structures. Examples of nonhydrogen bonded and cyclic structures are also described. The crystallographic results compare well with structural studies in solution, establishing that Aib peptides can provide rigid structural models for the development of spectroscopic methods of peptide conformational analysis.

Aminoisobutyric Acids↗

Conformational studies on host-guest peptides containing chiral alpha-methyl-alpha-amino acids. Comparison of the helix-inducing potential of alpha-aminoisobutyric acid, (S)-2-ethylalanine and (S)-2-methylserine.

The conformational behaviour of host-guest peptides of the type Ac-Ala-Xxx-Ala-Ala-Xxx-Ala-Ala-Xxx-Ala-Ala-NH-PEGM (Xxx = alpha-aminoisobutyric acid (Aib), (S)-2-ethylalanine ((S)-Iva), (S)-2-methylserine ((S)-alpha-MeSer)) has been studied by CD spectroscopy in CF3CH2OH, CH3OH, and water and by i.r. spectroscopy in CHCl3 and in the solid state. In this way the relative helix-inducing potential of the two chiral alpha-methyl-alpha-amino acids (S)-Iva and (S)-alpha-MeSer could be established in comparison to the strong helix-former Aib. The results show that (S)-Iva exerts a comparable helix-inducing effect as Aib, making this amino acid a valuable complementary tool for the stabilization or induction of helices. No significant helix-promoting effect was observed for (S)-alpha-MeSer in polar solvents; however, the i.r.-spectroscopic data in CHCl3 and in the solid state point to a helical conformation under these conditions. Possible reasons for the different behaviour of (S)-Iva and (S)-alpha-MeSer are briefly discussed.

Aminoisobutyric Acids↗

Sodium-stimulated active transport of aminoisobutyric acid by reconstituted vesicles from partially purified plasma membranes of mouse fibroblasts transformed by simian virus 40.

Plasma membrane fractions isolated from mouse fibroblast BALB/c 3T3 cells transformed by simian virus 40 were partially purified by treatment with dimethylmaleic anhydride followed by extraction with 2% cholate. The extracted proteins were combined with exogenous phospholipids and eluted through a Sephadex G50 column. Reconstituted vesicles thus obtained were shown to possess the ability of Na+-stimulated transport of alpha-aminoisobutyric acid. The simultaneous addition of NaSCN and alpha-aminoisobutyric acid to these vesicles produced a transient accumulation above the equilibrium level (overshoot, active transport). The Na+-stimulated transport of alpha-aminoisobutyric acid was sensitive to the accompanying anion and to the temperature of incubation. The results demonstrate that partially purified membrane proteins of mouse fibroblast cells can be incorporated into the liposomes that have the characteristics of Na+-stimulated and electrochemically sensitive active transport of alpha-aminoisobutyric acid.

Aminoisobutyric Acids↗

Distribution of alpha-aminoisobutyric acid in tissues of lean and genetically obese mice.

Distribution of tracer amounts of the nonmetabolizable neutral amino acid alpha-[1-14C]aminoisobutyric acid (AIB) between blood and several tissues was measured in lean and ob/ob mice over an 8-hr period. As AIB was injected on the basis of body weight and as ob/ob mice have a relatively low blood volume, absolute concentrations of AIB in blood and tissues were almost always higher in the obese than the lean mice. However, the ratio of AIB concentration in the tissues to that in the blood was clearly higher in skeletal muscle, diaphragm, and brain, and possibly higher in liver of the lean than of the obese animals. Ratios in heart were similar. The results suggest that lean and genetically obese mice differ in their capacity to transport amino acids between blood and various tissues.

Aminoisobutyric Acids↗

Stimulus-secretion coupling of arginine-induced insulin release. Effect of 2-aminoisobutyric acid upon ornithine decarboxylase activity and insulin secretion.

Recent studies have revealed that, in rat pancreatic islets, exogenous L-arginine or L-ornithine are converted to putrescine, spermidine and spermine. The possible role of polyamine generation in the secretory response of the B-cell to the cationic amino acids was investigated in islets either preincubated for 90 min with 2-aminoisobutyric acid (AIB; 10 mM) or removed from rats injected intraperitoneally with AIB (2.4 mmol) 210 min before sacrifice. Although AIB failed to exert any direct effect upon the secretory response of the islets to either L-arginine or L-ornithine, preincubation of the islets with AIB increased both the activity of ornithine decarboxylase and, to a modest extent, the insulinotropic action of L-arginine and L-ornithine. Likewise, after treatment with AIB in vivo, the activity of ornithine decarboxylase increased tenfold or more in kidney, liver, pancreas, parotid gland and islet homogenates. The secretory response to D-glucose was little affected in islets removed from AIB-treated rats, but the insulinotropic action of L-arginine and L-ornithine were slightly increased. These findings suggest that the de novo generation of polyamines only plays a restricted role in the secretory response of islet cells to L-arginine and L-ornithine.

Amino Acids↗

Infrequency of urinary excretion of beta-aminoisobutyric acid by healthy humans.

Urinary beta-aminoisobutyric acid (BAIB) levels were measured by modified ion-exchange chromatography. Daily BAIB levels are reported for a group of apparently healthy individuals, a subject with infective hepatitis and two human genetic variants who were high "excretors" of BAIB. The frequency of high "excretors" of BAIB was found using low-voltage paper electrophoresis. The frequency among 403 male European subjects was 5 (1.2%) and lower than previous estimates in the United Kingdom. The percentage recovery of fed BAIB suggests that Oriental "excretors" may have a different type of genetic lesion from non-Oriental "excretors". The clinical usefulness of urinary BAIB assays and qualitative screening is reviewed.

Adolescent↗

CD-n.m.r. study of the solution conformation of bradykinin analogs containing alpha-aminoisobutyric acid.

The conformation in aqueous solution of several alpha-aminoisobutyric acid (AIB)-containing analogs of bradykinin (BK) has been probed by complementary CD and 1H n.m.r. measurements. The conclusion reached is that substitution of AIB for Pro2 and/or Pro3 in BK stabilizes a degree of beta-turn conformation in the N-terminal tetrapeptide moiety of the resulting analogs. Changing the solvent from water to DMSO or TFE further enhances the contribution of particular hydrogen bonded structures to the time-averaged conformation of these peptides. Bradykinin and [AIB7]-BK adopt similar hydrogen bonded conformations in TFE, apparently with a contribution from a beta-turn involving their common Arg1-Pro2-Pro3-Gly4 moiety. The contrasting biological activities of BK and its AIB-analogs are considered in terms of the conformational analogy between the AIB-residue and cis' Pro and the propensity for a beta-turn at the N-terminus of the peptide.

Amino Acid Sequence↗

Insulin and exercise stimulate muscle alpha-aminoisobutyric acid transport by a Na+-K+-ATPase independent pathway.

Sodium ions are required for the active transport of amino acids such as alpha-aminoisobutyric acid (AIB) into skeletal muscle. To examine the role of Na+-K+-ATPase in this phenomenon, studies were carried out using the isolated perfused rat hindquarter preparation. Perfusion for 30 min with ouabain at a dose sufficient to inhibit the Na+-K+ pump (10(-4) M) inhibited the basal rate of AIB uptake in all muscles studied by up to 80%. However, it failed to inhibit the stimulation of AIB uptake, either by insulin (200 microU/ml) or electrically-induced muscle contractions. The increase in K+ release by the hindquarter in the presence of ouabain was the same under all conditions suggesting comparable inhibition of the Na+-K+ pump. These studies suggest that the basal, but not insulin or exercise-stimulated AIB transport into muscle is acutely dependent on a functional Na+-K+ pump. They also suggest that stimulated and basal uptake of AIB involve different mechanisms.

Aminoisobutyric Acids↗

Neutral amino acid transport in astrocytes: characterization of Na+-dependent and Na+-independent components of alpha-aminoisobutyric acid uptake.

Neutral amino acid transport is largely unexplored in astrocytes, although a role for these cells in blood-brain barrier function is suggested by their close apposition to cerebrovascular endothelium. This study examined the uptake into mouse astrocyte cultures of alpha-aminoisobutyric acid (AIB), a synthetic model substrate for Na+-dependent system A transport. Na+-dependent uptake of AIB was characteristic of system A in its pH sensitivity, kinetic properties, regulatory control, and pattern of analog inhibition. The rate of system A transport declined markedly with increasing age of the astrocyte cultures. There was an unexpectedly active Na+-independent component of AIB uptake that declined less markedly than system A transport as culture age increased. Although the saturability of the Na+-independent component and its pattern of analog inhibition were consistent with system L transport, the following properties deviated: (1) virtually complete inhibition of Na+-independent AIB uptake by characteristic L system substrates, suggesting unusually high affinity of the transporter; (2) apparent absence of trans-stimulation of AIB influx; (3) unusually concentrative uptake at steady state (the estimated distribution ratio for 0.2 mM AIB was 55); and (4) susceptibility to inhibition by N-ethylmaleimide. Direct study of the uptake of system L substrates in astrocytes is needed to confirm the present indications of high affinity and concentrative Na+-independent transport.

Amino Acids↗

Transport of alpha-aminoisobutyric acid across the blood-brain barrier studied with in situ perfusion of rat brain.

Transport of alpha-aminoisobutyric acid (AIB) from blood to brain in pentobarbital-anesthetized rats was examined using in situ perfusion. In situ perfusion with washed sheep red blood cells allowed the precise control of the composition of the perfusate that was necessary for a detailed examination of the transport of AIB. Retrograde perfusion at 4 ml/min through the left external carotid artery with oxygenated, artificial blood (hematocrit = 0.3) maintained a normal electroencephelogram during a 10 min experiment. The perfusate cerebral blood flow, at a value of 1.2 +/- 0.1 ml/g/min, and the perfusate cerebral plasma volume, at a value of 5.4 +/- 1.9 microliter/g, in the left frontal cortex were within the range of reported in vivo values. The in situ PS product for AIB (3.8 +/- 0.4 microliter/g/min) was higher than the value observed in vivo. AIB uptake was reduced to the in vivo value by 2 mM phenylalanine (1.3 +/- 0.3 microliter/g/min) and equally well by a mixture of neutral amino acids at their normal plasma concentrations but was unaffected by 2 mM methyl-AIB or removal of sodium from the perfusate. A kinetic analysis showed that the apparent Ki for phenylalanine inhibition of AIB transport was 19.8 +/- 4.9 microM. Thus, although AIB has affinity for the large neutral amino acid carrier in the blood-brain barrier, brain uptake by this mechanism in vivo is negligible due to competition by other amino acids in the plasma.

Amino Acids↗

Effects of dibutyryl cyclic AMP on the transport of alpha-methyl-D-glucoside and alpha-aminoisobutyric acid in separated tubules and brush border membranes from rabbit kidney.

The effect of dibutyryl cyclic AMP on the transport of alpha-methyl-D-glucoside and alpha-aminoisobutyric acid in separated tubules and purified brush border membranes from rabbit kidney was investigated using a rapid filtration procedure. Dibutyryl cyclic AMP stimulated the uptake of alpha-methyl-D-glucoside and alpha-aminoisobutyric acid by separated renal tubules in agreement iwth prior studies utilizing renal slices (Rea, C. and Segal, S. (1973) Biochim. Biophys. Acta 311, 615--624; Weiss, I.W., Morgan, K. and Phang, J.M. (1972) J. Biol. Chem. 247, 760--764). However, in contrast to previous reports, no preincubation of the tissue with dibutyryl cyclic AMP was required for stimulation of transport to be manifest. Dibutyryl cyclic AMP stimulated oxygen consumption by separated tubules suggesting that stimulation of transport may occur by a linkage with renal oxidative metabolism. Dibutyryl cyclic AMP increased the uptake of alpha-aminoisobutyric acid into purified renal brush border membranes. However the uptakes of alpha-methyl-D-glucoside, proline, leucine and phosphate into brush border membranes were significantly inhibited.

Aminoisobutyric Acids↗

Measurement of blood-brain barrier permeability of rats with alpha-aminoisobutyric acid during microdialysis: possible application to behavioral studies.

The purpose of this study was to determine the suitability of alpha-aminoisobutyric acid (AIB) as a marker of the integrity of the blood-brain barrier (BBB) during studies in which amino acid content in dialysates collected by microdialysis probes and behavior of the rat are studied concurrently. AIB (200 mg/kg) was injected intraperitoneally at 1800 h into 24-h fasted, ketamine-acepromazine anesthetized rats on either 10 or 30 days following guide cannula implantation. Dialysates from the medial preoptic area and blood from the tail vein were collected 1 h before and after the AIB injection. Analysis of amino acids, including AIB, in the collections was conducted by reverse-phase HPLC. In 21 of 24 rats, AIB in dialysates averaged less than 3% of plasma at 10, 30, and 50 min after AIB injection. In those rats unidirectional blood-to-brain transfer constant, K(i), of AIB was constant and a good relationship was found between dialysate amino acid concentrations and those predicted from calculations of transport based on the brain uptake index (BUI) for some amino acids. AIB concentration in dialysates was greater than 10% of plasma at 30 min in only 3 of 24 rats. We conclude that AIB can be used as a marker to monitor the integrity of the BBB during serial measurements of dialysate concentrations of amino acids and has application in behavioral studies.

Algorithms↗

Insulin-induced insulin resistance of alpha-aminoisobutyric acid transport in cultured human skin fibroblasts.

Cultured fibroblasts derived from skin biopsies were used to develop a system for studying insulin resistance in human tissue in vitro. Uptake of alpha-aminoisobutyric acid by cultured human skin fibroblasts was found to occur by a combination of saturable and nonsaturable processes. Insulin stimulated uptake by decreasing the Km of the saturable transport system from 0.58 mM to 0.26 mM. The maximal velocity of saturable uptake was 16.6 nmol/10(7) cells/min in both the presence and absence of insulin. Uptake of alpha-aminoisobutyric acid at 0.2 mM was studied in human skin fibroblasts with and without chronic exposure to insulin for 4 days at an initial concentration of 10 micrograms/ml. Unstimulated uptake was increased from 17 to 20 nmol/10(8) cells/min, and the increase in uptake due to maximal stimulation by insulin was unchanged at 16 nmol/10(8) cells/min in the cells exposed chronically to insulin. The apparent Km for insulin was increased from 80 microunits/ml to 2400 microunits/ml in the insulin-exposed cells. Thus, chronic exposure to insulin induces resistance of alpha-aminoisobutyric acid uptake by decreasing the apparent affinity for insulin.

Adult↗

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.

Amino Acids↗

Characteristics of alpha-aminoisobutyric acid transport in rat skeletal muscles.

alpha-Aminoisobutyric acid (AIB) transport into the intracellular compartment of extensor digitorum longus and soleus muscles was measured (in vitro) after allowance for the equilibration of the amino acid in the extracellular space. The latter was determined with three markers, [14C]inulin, 60Co-EDTA and [3H]mannitol. Net transport of AIB was subsequently divided into its two components, i.e. influx and efflux. Rates of influx were measured as the intracellular accumulation of [14C]AIB after a short incubation (5 min), and efflux was measured as the release of AIB with time (up to maximum of 50 min) from muscles that had previously been preloaded with AIB. This intracellular efflux was resolved into two phases, which probably represent two separate components of exit. The influence of extracellular Na+ on the transport of this neutral amino acid (representing the A system) was investigated. Na+ depletion resulted in lower accumulations of AIB, the effects becoming more pronounced with progressive depletions of external Na+. These changes arose from an inhibition of AIB influx, concomitant with an enhancement of its efflux. In contrast, all components of tyrosine transport (representing the L system) were unaffected by lowering external Na+ concentrations. The net accumulation of AIB was also suppressed by cortisol. This inhibitory effect was, however, Na+-dependent and resulted solely from the steroid's enhancement of AIB efflux, the hormone being without effect on AIB influx.

Aminoisobutyric Acids↗