Brain metabolism of branched-chain amino acids

Domestic swine (n=12 in each group) were randomized to daily treatment with the thromboxane A2 (TXA2) receptor antagonist BMS-180291 (group I), aspirin (group II), or no drug (group III) prior to prosthetic carotid graft implantation. Platelet and arterial wall receptor density were measured by equilibrium binding using 125I-BOP. At 6 weeks, means (s.e.m.) platelet receptor density (pmol/mg) had increased in groups I (3.3(0.6) versus 1.8(0.3); P<0.05) and II (2.6(0.6) versus 1.7(0.2); P<0.05), but not in group III (1.3(0.3) versus 1.2(0.2)). Aortic membrane TXA2 receptor density (fmol/mg) was significantly greater (P<0.05) in groups I (150(50)) and II (68(10)) compared with group III (39(6)). Chronic exposure to a TXA2 receptor antagonist or aspirin is associated with increased platelet and aortic receptor density in pigs.     

A study of branched-chain amino acid aminotransferase and isolation of mutations affecting the catabolism of branched-chain amino acids in Saccharomyces cerevisiae

The specific activity of branched-chain amino acid aminotransferase was highest when S. cerevisiae was grown in minimal medium containing a branched-chain amino acid as nitrogen source. Growth in complex media with glycerol or ethanol gave moderately high levels, whereas with glucose and fructose the specific activity was very low. Mutagenesis defined three genes (BAA1 to BAA3) required for branched-chain amino acid catabolism. The baa1 mutation reduced the specific activity of the aminotransferase, the stationary phase density in YEPD and caused gross morphological disturbance. Branched-chain amino acid aminotransferase is essential for sporulation.     

Ferrireductase from Pichia guilliermondii: properties and regulation of activity and synthesis

Iranian Jews represent an ancient community with a very high degree of inbreeding. Although the community remained relatively isolated, it had strong ties with Babylonian Jewry in Iraq. Several genetic disorders have been reported to be frequent among Iranian Jews, in particular, corticosterone methyloxydase deficiency type II, polyglandular syndrome, and rimmed vacuole myopathy. Based on the data collected in our clinic, recessive and dominant deafness also appear to be frequent. Other diseases, such as beta-thalassemia, achromatopsia, colobomatous microphthalmia, Dubin-Johnson syndrome, and congenital myasthenia gravis, were frequent in both the Iranian and Iraqi Jewish communities. The place of origin of the families within Iran and the results of molecular studies suggest some reason(s) for the high frequency of these disorders among Iranian Jews. While the high frequency of some of the disorders, such as corticosterone methyloxydase deficiency type II, represents a founder effect, in other diseases (such as beta-thalassemia) it was secondary to heterozygote advantage.     

The influence of bicarbonate supplementation on plasma levels of branched-chain amino acids in haemodialysis patients with metabolic acidosis

BACKGROUND: It has been hypothesized that correction of metabolic acidosis might improve the nutritional state of acidotic haemodialysis (HD) patients partly because of a reduced oxidation of branched-chain amino acids (BCAA). AIM: We investigated whether bicarbonate (Bic) supplementation in acidotic HD patients results in increased plasma levels of BCAA. METHODS: In a longitudinal study (run-in period, 2 months; study period, 6 months), the effect of Bic supplementation on plasma levels of BCAA was studied in 12 acidotic HD patients (7 men, 5 women, mean age 54 +/- 18 years) with a predialysis bicarbonate (Bic) concentration smaller or equal to 22 mmol/l. Bic was supplemented by increasing Bic concentration of the dialysate and by oral Bic supplementation. RESULTS: Predialysis Bic increased significantly during the study period (18.7 +/- 2.7 vs. 23.1 +/- 11.5 mmol/l). There was no change in nutritional parameters. However, plasma levels of the BCAA valine, leucine, and isoleucine increased significantly. CONCLUSIONS: In haemodialysis patients with metabolic acidosis, Bic supplementation over a 6-months period resulted in an increase in plasma levels of BCAA. Further study is needed to elucidate the mechanisms behind, and the clinical importance of the observed changes in plasma BCAA levels.     

Modulation of pentylenetetrazol-induced seizure activity by branched-chain amino acids and alpha-ketoisocaproate

Branched-chain amino acids, and mainly leucine act as nitrogen donors in the cerebral glutamate-glutamine cycle, thereby reducing brain excitability. Rats equipped with cortical electrodes received 300 mg/kg of leucine, isoleucine, valine or the ketoacid of leucine, alpha-ketoisocaproate at 2 h before the induction of seizures by 40 mg/kg pentylenetetrazol. Control groups received saline or a commercial mixture of amino acids, Vamine(R). Leucine and isoleucine increased the latency to absence-like and tonic-clonic seizures but did not influence the duration of the tonic-clonic seizure. Vamine(R), valine and alpha-ketoisocaproate had no effect. These data are consistent with the role of leucine in buffering brain glutamate concentration.     

Oral branched-chain amino acids do not improve exercise capacity in McArdle disease

To determine whether oral branched-chain amino acids (BCAAs) improve exercise capacity, six fasting patients with McArdle's disease were given a solution of BCAA (77 mg/kg) or a control noncaloric beverage 30 minutes before exercise. The BCAA meal tripled plasma BCAA levels, increased BCAA catabolism as indicated by greater exercise increases in plasma glutamine and alanine, but lowered mean peak free fatty acid levels and reduced exercise capacity in five of six patients. Lower work capacity may be attributed to a net reduction in muscle fuel availability after BCAA administration.     

Biocontrol of mold growth in high-moisture wheat stored under airtight conditions by Pichia anomala, Pichia guilliermondii, and Saccharomyces cerevisiae

Pichia anomala inhibits the growth of Penicillium roqueforti and Aspergillus candidus on agar. In this investigation, antagonistic activity on agar against 17 mold species was determined. The abilities of Pichia anomala, Pichia guilliermondii, and Saccharomyces cerevisiae to inhibit the growth of the mold Penicillium roqueforti in nonsterile high-moisture wheat were compared by adding 10(3) Penicillium roqueforti spores and different amounts of yeast cells per gram of wheat. Inoculated grain was packed in glass tubes, incubated at 25 degrees C with a restricted air supply, and the numbers of yeast and mold CFU were determined on selective media after 7 and 14 days. Pichia anomala reduced growth on agar plates for all of the mold species tested in a dose-dependent manner. Aspergillus fumigatus and Eurotium amstelodami were the most sensitive, while Penicillium italicum and Penicillium digitatum were the most resistant. Pichia anomala had the strongest antagonistic activity in wheat, with 10(5) and 10(6) CFU/g completely inhibiting the growth of Penicillium roqueforti. Inhibition was least pronounced at the optimum temperature (21 degrees C) and water activity (0.95) for the growth of Penicillium roqueforti. Pichia guilliermondii slightly reduced the growth of Penicillium roqueforti in wheat inoculated with 10(5) and 10(6) yeast CFU/g. S. cerevisiae inhibited mold growth only weakly at the highest inoculum level. Pichia anomala grew from 10(3) to 10(7) CFU/g of wheat in 1 week. To reach the same level, Pichia guilliermondii had to be inoculated at 10(4) CFU while S. cerevisiae required an inoculum of 10(5) CFU to reach 10(7) CFU/g of wheat.     

Regulation of glutamate dehydrogenase by branched-chain amino acids in skeletal muscle from rats and chicks

Neurotrophin-4 (NT-4) is a member of a family of neurotrophic factors, the neurotrophins, that control survival and differentiation of vertebrate neurons (2-4). Besides being the most recently discovered neurotrophin in mammals, and the least well understood, several aspects distinguish NT-4 from other members of the neurotrophin family. It is the most divergent member and, in contrast to the other neurotrophins, its expression is ubiquitous and appears to be less influenced by environmental signals. NT-4 seems to have the unique requirement of binding to the low-affinity neurotrophin receptor (p75LNGFR) for efficient signalling and retrograde transport in neurons. Moreover, while all other neurotrophin knock-outs have proven lethal during early postnatal development, mice deficient in NT-4 have so far only shown minor cellular deficits and develop normally to adulthood. Is NT-4 a recent addition to the neurotrophic factor repertoire in search of a crucial function, or is it an evolutionary relic, a kind of wisdom tooth of the neurotrophin family?     

A flavinogenic mutant of the yeast Pichia guilliermondii with impaired iron transport

We recently demonstrated that net fluid uptake occurs in the capillary system of the inner medulla. To define the site of fluid uptake, the concentration of protein was determined in plasma from descending vasa recta at the base and tip of the exposed papilla in Munich-Wister rats. The vasa recta plasma-to-arterial plasma protein concentration ratio (VR/P) was 1.43 +/- 0.09 at the base and 1.66 +/- 0.09 at the tip. These results, which indicate fluid loss from the descending vasa recta, are difficult to explain on the basic of hydraulic and oncotic forces alone. The osmolality of the contents of descending vasa recta increased between base and tip (delta = 72 +/- 30 mosmol/kg H2O). If the increase in osmolality of plasma in descending vasa recta lags behind that of the adjacent medullary interstitium, a transcapillary osmotic driving force exists favoring water loss from descending vessels. It is concluded that fluid uptake by the inner medullary circulation occurs beyond descending vasa recta in interconnecting capillaries or ascending vasa recta. In our view the most likely interpretation of these results is that fluid movement across vasa recta in the inner medulla is influenced by three forces: those owing to transcapillary differences in osmotic, oncotic, and hydraulic pressures.     

Regulation of branched-chain amino acid metabolism in the lactating rat

There is evidence that during lactation, uptake of the essential branched-chain amino acids (BCAA) by mammary glands exceeds their output in milk protein. In this study, we have measured the potential of lactating rats to catabolize BCAA. The activity, relative protein and specific mRNA levels of the first two enzymes in the BCAA catabolic pathway, branched-chain aminotransferase (BCAT) and branched-chain alpha-keto acid dehydrogenase (BCKD), were measured in mammary gland, liver and skeletal muscle obtained from rat dams at peak lactation (12 d), from rat dams 24 h after weaning at peak lactation and from age-matched virgin controls. Western analysis showed that the mitochondrial BCATm isoenzyme was found in mammary gland. Comparison of lactating and control rats revealed that tissue BCATm activity, protein and mRNA were at least 10-fold higher in mammary tissue during lactation. Values were 1.3- to 1. 9-fold higher after 24 h of weaning. In mammary gland of lactating rats, the BCKD complex was fully active. In virgin controls and weaning dams, only about 20% of the complex was in the active state. Hypertrophy of the liver and mammary gland during lactation resulted in a 73% increase in total oxidative capacity in lactating rats. The results are consistent with increased expression of the BCATm gene in the mammary gland during lactation, whereas oxidation appears to be regulated primarily by changes in activity state (phosphorylation state) of BCKD.     

The antinociceptive effects of branched-chain amino acids: evidence for their ability to potentiate morphine analgesia

The effect of branched-chain amino acids (BCAA) on pain threshold was studied in rats. Nociception was induced by the hot-plate analgesia meter, a method measuring supraspinally organized pain responses. After a single intravenous injection of BCAA (320 mg/kg), the percent change in latency time to the pain response significantly increased by 19% in 60 min, and by 22% in 75 min (p < 0.005), as compared to an injection of an equal volume of a standard concentration of an amino acid solution or physiological saline. Subsequently, we studied the interaction of BCAA with opioid-type analgesia. In combination with intravenously injected morphine (3 mg/kg), BCAA significantly potentiated and prolonged the action of morphine using the hot-plate test. From 5 min after morphine injection, the latencies to a pain response were markedly higher with the combination of BCAA and morphine (+80% and +89% at 5 min after morphine injection, if BCAA was administered 45 or 60 min prior to morphine injection, respectively) when compared with the effect of morphine alone (+13% at 5 min; p < 0.005). BCAA demonstrated analgesic effects, which, in combination with morphine, potentiated and prolonged the antinociceptive action of morphine. BCAA may represent a new adjunct treatment modality for acute and chronic pain, and give us further insight into the mechanisms of pain control.     

Performance of chicks fed diets formulated to minimize excess levels of essential amino acids

Studies were conducted in which levels of essential amino acids in excess of the minimum requirements were minimized in broiler diets composed of commercially available feed stuffs and synthetic amino acid supplements. Growth rate and efficiency of feed utilization of chicks fed such diets were equal to that attained by chicks fed diets formulated by conventional means when grown under conditions where heat stress was not a factor and significantly improved when fed under conditions of heat stress. Improvements were obtained in efficiency of protein and calorie utilization using this technique of formulation.     

Alterations in K+ evoked profiles of neurotransmitter and neuromodulator amino acids after focal ischemia-reperfusion

e present evidence that nestmate discrimination in the eusocial paper wasp, Polistes dominulus, is context dependent. We compared aggression levels between nestmates and non-nestmates in dyads consisting of a pair of either nestmates or non-nestmates, and triads consisting of either three nestmates, three non-nestmates, or two nestmates and a non-nestmate. In 130 of the 237 total trials, a nest fragment (containing both brood and eggs) from the nest of some, all or none of the interactants was placed into the interaction arena. Polistes dominulus workers recognized and discriminated nestmates from non-nestmates, familiar from unfamiliar nest material and neighbours from non-neighbours. These findings suggest that nestmate and neighbour discrimination are context dependent: discrimination occurs when either the presence of a nestmate or a familiar nest fragment indicate the proximity of the colony. The context-dependent variation in aggression levels is best described by multiple, context-dependent shifts in an acceptance threshold. Thus this study provides the most extensive, critical support yet obtained for Reeve's (1989, American Naturalist, 133, 407-435) optimal acceptance threshold model. Copyright 1998 The Association for the Study of Animal Behaviour     

Responses of circulating urea cycle and branched-chain amino acids to feeding in adult and aged Fischer-344 rats

The goal of our study was to identify cytokine combinations that would result in simultaneous ex vivo expansion of both the megakaryocyte (Mk) and granulocyte lineages, since these cell types have the potential to reduce the periods of thrombocytopenia and neutropenia following chemotherapy. We investigated the effects of cytokine combinations on expansion of the Mk (CD41a+ cells and colony forming unit [CFU]-Mk) and granulocyte (CD15+ cells and CFU-granulocyte/monocyte [GM]) lineages. Peripheral blood CD34+ cells were cultured in serum-free medium with interleukin 3 (IL-3), stem cell factor (SCF), and various combinations of thrombopoietin (TPO), IL-6, GM-CSF, and/or G-CSF. The Mk lineage was primarily influenced by TPO in our cultures, although Mk and CFU-Mk numbers were increased when TPO was combined with IL-6. The primary stimulator of the granulocyte lineage was G-CSF, although many synergistic and additive effects were observed with addition of other factors. Expansion of CFU-GM increased upon addition of more cytokines. The cytokine combination of IL-3, SCF, TPO, IL-6, GM-CSF and G-CSF produced the greatest number of granulocytes and CFU-GM. The minimum cytokines necessary for expansion of both the Mk and granulocyte lineages included TPO and G-CSF, since no other factors examined could increase Mk and granulocyte numbers to the same extent. The number of hematopoietic progenitors produced in our culture system should be sufficient for successful engraftment following myelosuppressive therapy if produced on a scale of about one liter.     

The influence of intravenous infusions of glucose and amino acids of pancreatic enzyme and mucosal protein synthesis in human subjects

BACKGROUND: Animal studies have shown that the synthesis and secretion of pancreatic enzymes and the turnover of mucosal proteins is strongly influenced by diet. METHODS: To determine whether the absorbed products of digestion are responsible for these changes, we investigated in groups of five healthy volunteers, the effects of i.v. infusions of amino acids (0.08 g/kg/h) and glucose (0.3 g/kg/h) on pancreatic enzyme and mucosal protein synthesis. Proteins were labeled in vivo by a 4-hours i.v. infusion of 14C-leucine and the enteric infusion of 3H-leucine tracer, during simultaneous cholecystokinin stimulation and duodenal collection of secreted pancreatic enzymes. Labeling of mucosal proteins was measured by endoscopic biopsy. RESULTS: The amino acid infusions elevated plasma amino acid levels, and the glucose infusions increased both glucose and insulin concentrations. The rates for amylase and trypsin secretion were significantly lower during the first 2 hours of glucose infusion and the rate of synthesis of trypsin was delayed by i.v. amino acid infusions from 52.1 +/- 4.1 to 77.6 +/- 8.5 minutes. Mucosal protein turnover rates were unaffected. 3H-labeling via the enteral route showed similar enzyme synthesis rates but variable mucosal incorporation rates. CONCLUSIONS: I.v. nutrients do not appear to stimulate the synthesis of pancreatic and mucosal proteins in human subjects.     

Stimulation of albumin synthesis by keto analogues of amino acids

(1) The effect of ketoanalogues of branched-chain amino acids on albumin synthesis was examined in two biological systems using the [14C]carbonate technique. (2)alpha-Ketocaproic acid, the ketoanalogue of leucine, was able to reverse the reduced synthesis rate observed when isolated livers, from well-nourished animals were perfused with blood from rats deprived of dietary protein for 48 h. (3) A mixture of ketoanalogues of the three branched-chain amino acids, leucine, isoleucine and valine, was able to increase albumin synthesis per unit dry liver weight to above normal levels when administered intragastrically to rats 16 h after partial hepatectomy.     

Effect of exercise intensity on free tryptophan to branched-chain amino acids ratio and plasma prolactin during endurance exercise

The potential of exercise-induced changes in peripheral amino acids to alter blood prolactin levels through a serotonergic system modification was investigated in 8 male athletes. In two trials, subjects (N = 8) exercised on a cycle ergometer for 5 hr. The intensity of exercise corresponded to 55% VO2max (T55) or 75% VO2max (T75), respectively. In each trial, each subject received a 25-g energy bar (111 kcal) every 60 min, as well as 300 ml of a 6% carbohydrate solution (90 kcal) every 30 min of exercise duration. Plasma glucose and insulin declined (p < or = .05) in both trials during exercise. Ammonia was augmented (p < or = .05) above the baseline concentration after 120 min in both trials. During the last 2 hr of exercise, plasma free fatty acids were higher (p < or = .05) in T75 than in T55. During this time, the plasma free TRP/BCAA ratio was also augmented (p < or = .05) in T75, while no change was induced in T55. Plasma prolactin did not change in T55, while an increase (p < or = .05) was found in T75. The findings may further support the hypothesis that during endurance exercise changes in peripheral amino acid concentration may influence prolactin response via serotonergic system modifications.     

Role of excitatory amino acids in the regulation of dopamine synthesis and release in the neostriatum

We have explored the role of excitatory amino acids in the increased dopamine (DA) release that occurs in the neostriatum during stress-induced behavioral activation. Studies were performed in awake, freely moving rats, using in vivo microdialysis. Extracellular DA was used as a measure of DA release; extracellular 3,4-dihydroxyphenylalanine (DOPA) after inhibition of DOPA decarboxylase provided a measure of apparent DA synthesis. Mild stress increased the synthesis and release of DA in striatum. DA synthesis and release also were enhanced by the intra-striatal infusion of N-methyl-D-aspartate (NMDA), an agonist at NMDA receptors, and kainic acid, an agonist at the DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate (AMPA)/kainate site. Stress-induced increase in DA synthesis was attenuated by co-infusion of 2-amino-5-phosphonovalerate (APV) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), antagonists of NMDA and AMPA/kainate receptors, respectively. In contrast, intrastriatal APV, CNQX, or kynurenic acid (a non-selective ionotropic glutamate receptor antagonist) did not block the stress-induced increase in DA release. Stress-induced increase in DA release was, however, blocked by administration of tetrodotoxin along the nigrostriatal DA projection. It also was attenuated when APV was infused into substantia nigra. Thus, glutamate may act via ionotropic receptors within striatum to regulate DA synthesis, whereas glutamate may influence DA release via an action on receptors in substantia nigra. However, our method for monitoring DA synthesis lowers extracellular DA and this may permit the appearance of an intra-striatal glutamatergic influence by reducing a local inhibitory influence of DA. If so, under conditions of low extracellular DA glutamate may influence DA release, as well as DA synthesis, by an intrastriatal action. Such conditions might occur during prolonged severe stress and/or DA neuron degeneration. These results may have implications for the impact of glutamate antagonists on the ability of patients with Parkinson's disease to tolerate stress.     

Utilization of methylmalonate for the synthesis of branched-chain fatty acids by preparations of chicken liver and sheep adipose tissue

1. The utilization of methyl[2-14C]malonyl-CoA for fatty acid synthesis was investigated using synthetase preparations from chicken liver and sheep adipose tissue. 2. The rate of fatty acid synthesis from acetyl-CoA and malonyl-CoA was greatly diminished in the presence of methylmalonyl-CoA. 3. In the absence of malonyl-CoA, methylmalonyl-CoA was utilized for fatty acid synthesis only very slowly by the synthetase from sheep adipose tissue and not at all by that from chicken liver. 4. Despite the inhibitory effect of methylmalonyl-CoA on fatty acid synthesis from malonyl-CoA, it was utilized by the synthetase preparations from both species to produce a complex mixture of methyl-branched fatty acids.