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Mechanisms of ammonia and ammonium ion toxicity in animal cells: transport across cell membranes.

A model for transport of ammonia and ammonium ions across cell membranes is presented. The model suggests that ammonium ions compete with potassium ions for inward transport, over the cytoplasmic membrane, via potassium transport proteins like the Na+/K(+)-ATPase and the Na+K+2Cl(-)-cotransporter. It also explains the difference between the ammonia/ammonium that is added to the cells and which is formed by the cells during metabolism of amino acids, especially glutamine and glutamate. The ammonium transport and subsequent events lead to predictable intracellular and extracellular pH (pHe) changes. Experiments which verified the model and the predicted consequences were performed by measurements of the pHe in concentrated cell suspensions. Addition of ammonium ions caused a time-dependent pHe increase which was inhibited by potassium ions. The test system is not per se specific for transport measurements but the effect of potassium ions on the pHe strongly favors our suggested model. Simple diffusion of ammonium ions would not be counteracted by potassium ions. The results show that ammonium ion transport in the murine myeloma cell line (Sp2/0-Ag14) used is inhibited by an excess of potassium ions. Results from experiments with specific inhibitors of suggested transport proteins were not conclusive. It is postulated that one important toxic effect of ammonia/ammonium is an increased demand for maintenance energy, caused by the need to maintain ion gradients over the cytoplasmic membrane. The results also suggest that potassium ions can be used to detoxify ammonia/ammonium in animal cell cultivations.

Amiloride↗

A model for the study of Helicobacter pylori interaction with human gastric acid secretion.

We present a comprehensive mathematical model describing Helicobacter pylori interaction with the human gastric acid secretion system. We use the model to explore host and bacterial conditions that allow persistent infection to develop and be maintained. Our results show that upon colonization, there is a transient period (day 1-20 post-infection) prior to the establishment of persistence. During this period, changes to host gastric physiology occur including elevations in positive effectors of acid secretion (such as gastrin and histamine). This is promoted by reduced somatostatin levels, an inhibitor of acid release. We suggest that these changes comprise compensatory mechanisms aimed at restoring acid to pre-infection levels. We also show that ammonia produced by bacteria sufficiently buffers acid promoting bacteria survival and growth.

Ammonia↗

Mechanisms of NH4+ and NH3 transport during hypokalemia.

Along the collecting duct, secretion of ammonium (NH) is thought to occur through active H+ secretion in parallel with the non-ionic diffusion of ammonia (NH3). Thus NH3 is secreted into the collecting duct lumen down its concentration gradient. Moreover, the low NH permeability and high NH3 permeability observed in collecting duct epithelia minimizes back diffusion of NH. In general, an increase in the NH3 concentration gradient between the interstitium and the collecting duct lumen correlates with increased NH secretion. However, our laboratory and others have shown an important role of direct NH transport by the Na,K-ATPase. As K+ and NH compete for a common extracellular binding site on the Na,K-ATPase, reduced interstitial K+ concentration, such as during hypokalemia, augments NH uptake. Na,K-ATPase-mediated NH uptake provides an important source of H+ for net acid secretion during hypokalemia and contributes to the increase in NH excretion and metabolic alkalosis observed in this treatment model.

Ammonia↗

Differential regulation of three functional ammonium transporter genes by nitrogen in root hairs and by light in leaves of tomato.

To elucidate the role of NH4+ transporters in N nutrition of tomato, two new NH4+ transporter genes were isolated from cDNA libraries of root hairs or leaves of tomato. While LeAMT1;2 is closely related to LeAMT1;1 (75.6% amino acid identity), LeAMT1;3 is more distantly related (62.8% identity) and possesses two short upstream open reading frames in the 5' end of the mRNA and a particularly short N-terminus of the protein as unique features. When expressed in yeast mutants defective in NH4+ uptake, all three genes complemented NH4+ uptake. In roots of hydroponically grown plants, transcript levels of LeAMT1;2 increased after NH4+ or NO3- supply, while LeAMT1;1 was induced by N deficiency coinciding with low glutamine concentrations, and LeAMT1;3 was not detected. In aeroponic culture, expression of LeAMT1;1 and LeAMT1;2 was higher in root hairs than in the remaining root fraction. Growth of plants at elevated CO2 slightly decreased expression of LeAMT1;2 and LeAMT1;3 in leaves, but strongly repressed transcript levels of chloroplast glutamine synthetase and photorespiratory serine hydroxymethyl-transferase. Expression of LeAMT1;2 and LeAMT1;3 showed a reciprocal diurnal regulation with highest transcript levels of LeAMT1;3 in darkness and highest levels of LeAMT1;2 after onset of light. These results indicate that in tomato at least two high-affinity NH4+ transporters, LeAMT1;1 and LeAMT1;2, are differentially regulated by N and contribute to root hair-mediated NH4+ acquisition from the rhizosphere. In leaves, the reciprocally expressed transporters LeAMT1;2 and LeAMT1;3 are supposed to play different roles in N metabolism, NH4+ uptake and/or NH3 retrieval during photorespiration.

Amino Acid Sequence↗

Mutation in ntrC gene leading to the derepression of nitrogenase synthesis in Rhodobacter sphaeroides.

The Rhodobacter sphaeroides mutants Drn12 and Drn21 derepressed for nitrogenase synthesis in the presence of ammonia and impaired in utilization of certain nitrogen sources have been analyzed. Both mutants show a low level of expression of the glnBA operon. The DNA fragment restoring the wild-type phenotype to these mutants contains the 3'-portion of ntrB gene and the entire ntrC gene. Sequence analysis showed that Drn12 bears a missense mutation in the ntrC gene. The mutation results in the replacement of a glycine residue by aspartate within the N-terminal domain of the NtrC protein. Pleiotropic phenotypes of Drn12 and Drn21 appear to be associated with an alteration in the regulation of glnBA expression.

Bacterial Proteins↗

Increased ammoniagenesis and the renal tubular effects of potassium depletion.

The cause of the morphological changes and functional defects in the renal tubule seen in patients with severe potassium depletion is unknown. In man and animals potassium status is a major factor regulating ammonia synthesis in the kidney and urinary ammonium excretion. A primary effect of potassium depletion is to cause an increase in ammoniagenesis by the renal tubular cells. It is proposed that the vacuolation of the renal tubular cells and the functional defects of tubular proteinuria, polyuria, resistance to arginine vasopressin, renal resistance to the action of parathyroid hormone, and increased urinary excretion of N-acetyl-beta-glucosaminidase found in potassium depletion are secondary effects caused by high concentrations of ammonia in the renal tubular cells.

Acetylglucosaminidase↗

Leaf age-related differences in apoplastic NH(4)(+) concentration, pH and the NH(3) compensation point for a wild perennial.

Extracts of the foliar apoplast of leaves of different ages of Luzula sylvatica (Huds.) Gaud. were prepared by vacuum infiltration and centrifugation. Measurements of pH and concentration were performed on extracts. From these bioassay measurements the relative magnitude of NH(3) compensation points for leaves of different ages were inferred. Young leaves were found to have much higher apoplast pH than old leaves, leading to the calculation of 4-10-fold higher NH(3) compensation points. Such age-related differences in the NH(3) compensation point are considerably larger than those previously reported. Apoplast pH and concentration were found to increase during leaf expansion before declining prior to senescence. Bulk foliar tissue pH, and total N concentrations were also found to be generally higher in young leaves than in old leaves. Where a significant correlation was found, total foliar N, bulk tissue foliar and the calculated NH(3) compensation point were all found to increase with N supplied to roots, whilst apoplast and bulk tissue H(+) concentrations were found to decline. The potential of bulk foliar tissue measurements to act as simple predictors of the NH(3) compensation point is discussed.

Ammonia↗