Effect of Inhibition of Gluconeogenesis on Ammonia Production in the Perfused Rat Kidney

1976 ◽  
Vol 50 (6) ◽  
pp. 493-498 ◽  
Author(s):  
B. D. Ross
Keyword(s):  
Rat Kidney ◽  
Glucose Production ◽  
Ammonia Production ◽  
Primary Event ◽  
Perfused Rat Kidney ◽  
Lactate Uptake ◽  
Ammonia Formation ◽  
Glutamine Uptake

1. Gluconeogenesis from lactate or from glutamine is inhibited by 90–100% by sodium quinolinate (1 mmol/l) or 3-mercaptopicolinate (150 nmol/l) in the perfused rat kidney. l-Tryptophan is not metabolized and is without effect. 2. Lactate uptake and glucose production are inhibited to the same degree by 3-mercaptopicolinate in the kidneys of well-fed or starved rats. 3. Inhibition of gluconeogenesis from glutamine (1 mmol/l) by 3-mercaptopicolinate is accompanied by 50% inhibition of ammonia production, and 34% inhibition of glutamine uptake, in the kidneys of acidotic rats. Ammonia production from glutamine was not inhibited in kidneys from non-acidotic rats. 4. It is concluded that the increased rate of gluconeogenesis from glutamine which occurs in acidotic rats is an essential and primary event regulating all of the increase in ammonia formation.

Ammonia production and glutamine incorporation into glutathione in the functioning rat kidney

1979 ◽  
Vol 57 (3) ◽  
pp. 233-237 ◽  
Author(s):  
T. C. Welbourne
Keyword(s):  
Resting State ◽  
Loading Condition ◽  
Rat Kidney ◽  
Incorporation Rate ◽  
Ammonia Production ◽  
Glutamine Concentration ◽  
Glutathione Synthesis ◽  
Isolated Perfused Rat Kidney ◽  
Perfused Rat Kidney ◽  
Glutamine Uptake

Incorporation of glutamine's γ-glutamyl moiety into glutathione was studied in the isolated perfused rat kidney under two conditions: (1) a resting state and (2) during reabsorption of a filtered glycylglycine load (designed to stimulate glutathione breakdown). Glutamine uptake and ammonia production were monitored simultaneously with [14C]glutamine (γ-glutamyl moiety) incorporation into glutathione.Renal glutathione content was highly dependent upon the presence of glutamine; in its absence, glutathione levels fell 40% in 30 min indicating synthesis does not keep pace with degradation. Presenting the kidney with a glycylglycine load doubled the rate of glutathione degradation. However, perfusing kidneys with physiological L-glutamine concentration (1 mM) maintained glutathione levels significantly above those in glutamine's absence in either condition 1 or 2. Evidence that glutamine functions as the γ-glutamyl donor in glutathione synthesis was established by the significant incorporation of [14C]glutamine into glutathione during condition 1 and the accelerated incorporation rate during glycylglycine loading (condition 2). Stimulating glutathione breakdown (condition 2) resulted in a significant increase in renal glutamine uptake and ammonia production apparently via the γ-glutamyltransferase pathway since inhibiting this activity blocked the incremental rise in glutamine uptake during glycylglycine loading. These results are consistent with the interpretation that stimulating γ-glutamyl cycle activity with a γ-glutamyl acceptor (glycylglycine) accelerates γ-glutamyl donor uptake (glutamine) and ammonia production via the γ-glutamyltransferase pathway.

Mithcondrial glutamine permeability and renal ammonia production in metabolic acidosis

1980 ◽  
Vol 239 (1) ◽  
pp. E51-E56
Author(s):  
T. C. Welbourne ◽  
G. T. Bazer
Keyword(s):  
Mitochondrial Membrane ◽  
Rat Kidney ◽  
Water Volume ◽  
Reflection Coefficients ◽  
Basal Region ◽  
Ammonia Production ◽  
Isolated Mitochondria ◽  
Membrane Feeding ◽  
Glutamine Uptake

These experiments were undertaken to determine the correspondence between acidosis-induced in situ motochondrial glutamine uptake and the process by which glutamine moves across the mitochondrial membrane. Feeding rats 1.5% NH4Cl for 2 wk accelerated the in situ uptake rate from 0.12 +/- 0.08 to 1.89 +/- 0.28 mu mol/min or some 16-fold. To determine glutamine uptake independent of its metabolic conversion, D-glutamine was employed. In isolated mitochondria from non-acidotic rat kidneys, D-glutamine diffused into 71 +/- 10% of the mitochondria water volume; in acidotic mitochondria the diffusion volume increased to 127 +/- 16%. The reflection coefficients (sigma) for a series of amides, including glutamine, were determined by gravimetrically following volume decrements in increasing concentrations of solute; D-glutamine's sigma fell from 1.05 +/- 0.08 to 0.50 +/- 0.06 in acidotic rat kidney mitchondria, The permeability coefficients corresponding to the measured sigma were 10(-7) cm/s and 10(-5) cm/s in nonacidosis and acidosis, respectively. When viewed in situ proximal tubule mitochondria undergo dramatic alterations during chronic acidosis. These involved an enlargement in the mitochondria particularly in the basal region of the cell and a reduction in number. Furthermore, numerous autophagic vacuoles, containing mitochondria, appear in the basal region. The findings are consistent availability that becomes activated during acidosis as a consequence of mitochondrial glutamine permeability resulting in increased ammoniagenesis and accelerated organelle turnover.

Ammoniagenesis: D-Glutamyltransferase as a Source of Ammonia in the Rat Kidney

1975 ◽  
Vol 53 (8) ◽  
pp. 930-933 ◽  
Author(s):  
P. Wadoux ◽  
T. C. Welbourne
Keyword(s):  
Glutamate Dehydrogenase ◽  
Rat Kidney ◽  
Ammonia Production ◽  
Normal Kidney ◽  
Isolated Perfused Rat Kidney ◽  
Perfused Rat Kidney ◽  
Total Ammonia ◽  
Normal Rat

The contribution of D-glutamyltransferase (D-GT) (EC 2.3.2.1) to total renal ammonia production was determined by employing DL-methionine-DL-sulfoximine (MSO) as an inhibitor of D-GT. Rat kidney homogenates were assayed for NH3-liberating activity under optimal D-GT or γ-glutamyltranspeptidase (γ-GTP) (EC 2.3.2.2) conditions. MSO inhibits only D-GT activity. The contribution of D-GT to total renal ammonia production was then evaluated in the isolated perfused rat kidney employing identical substrate (5 mML-glutamine) and inhibitor (15 mM MSO) concentrations as employed in the homogenate study. Under these conditions, MSO inhibits 70% of the total ammonia production by the normal kidney; in addition, the ratio of ammonia produced per glutamine taken up rose from 1.0 to 1.8. In kidneys from chronically acidotic rats, MSO reduced total ammonia production only 35% while the NH3/glutamine ratio rose from 1.0 to 1.8. D-GT appears to be the predominant source of NH3 production in the normal rat kidney; γ-GTP does not contribute significantly. The rise in the NH3/glutamine ratio after D-GT inhibition is consistent with glutamine utilization via the activated mitochondrial glutaminase (EC 3.5.1.2) – glutamate dehydrogenase (EC 1.4.1.2) pathway.

Glutamine synthesis in the perfused rat kidney and in isolated rat cortical tubules: regulation by pH

1985 ◽  
Vol 69 (6) ◽  
pp. 701-707 ◽  
Author(s):  
Henri Lanctin ◽  
John T. Brosnan ◽  
Brian D. Ross
Keyword(s):  
Metabolic Acidosis ◽  
Isotope Dilution ◽  
Rat Kidney ◽  
Ammonia Excretion ◽  
Ammonia Production ◽  
Dilution Technique ◽  
Isotope Dilution Technique ◽  
Perfused Rat Kidney ◽  
Glutamine Synthesis ◽  
Isolated Rat

1. The possible involvement of renal glutamine synthesis in the regulation of ammoniagenesis during metabolic acidosis has been investigated. 2. Using an isotope dilution technique glutamine synthesis has been demonstrated in the intact perfused rat kidney, despite zero net balance for glutamine in this preparation. 3. Inhibition of glutamine synthesis resulted in increased ammonia production in isolated cortical tubules. The rates of these processes were comparable. 4. We conclude that recycling of nitrogen through glutamine synthesis and glutamine hydrolysis could play a quantitatively significant role in the control of ammonia excretion by the kidney, particularly in acute acidosis where the fall in pH seems to inhibit glutamine synthesis.

scholarly journals Serine synthesis by an isolated perfused rat kidney preparation

1985 ◽  
Vol 230 (2) ◽  
pp. 303-311 ◽  
Author(s):  
R C Scaduto ◽  
E J Davis
Keyword(s):  
Aspartate Aminotransferase ◽  
Essential Role ◽  
Rat Kidney ◽  
Glucose Production ◽  
Complete Inhibition ◽  
Maximal Rate ◽  
Isolated Perfused Rat Kidney ◽  
Perfused Rat Kidney ◽  
Glucose Synthesis

The isolated perfused rat kidney was shown to synthesize serine from aspartate or glutamate, both of which are also precursors of glucose. The major products of aspartate metabolism were ammonia, serine, glutamate, glucose, glutamine and CO2. Perfusion of kidneys with aspartate in the presence of amino-oxyacetate resulted in a near-complete inhibition of aspartate metabolism, illustrating the essential role of aspartate aminotransferase in the metabolism of this substrate. Radioactivity from 14C-labelled aspartate and from 14C-labelled glycerol was incorporated into serine and glucose. Production of both glucose and serine from aspartate was suppressed in the presence of 3-mercaptopicolinic acid. These data provide evidence for the operation of the phosphorylated and/or non-phosphorylated pathway for serine production to the presence of 3-mercaptopicolinic acid. This is explained by simultaneous glycolysis. The rate of glucose production, but not that of serine, was greater in kidneys perfused with glutamate or with aspartate plus glycerol than the rates obtained by perfusion with aspartate alone. These data are taken to suggest that serine synthesis occurred at a near-maximal rate, and that the capacity of the kidney for serine synthesis from glucose precursors is lower than that for glucose synthesis.

Effect of acute metabolic acidosis on ammonia metabolism in kidney

1989 ◽  
Vol 256 (2) ◽  
pp. F321-F328
Author(s):  
R. T. Bogusky ◽  
R. L. Dietrich
Keyword(s):  
Metabolic Acidosis ◽  
Glutamate Dehydrogenase ◽  
Rat Kidney ◽  
Methionine Sulfoximine ◽  
Amide Nitrogen ◽  
Ammonia Metabolism ◽  
Perfused Rat Kidney ◽  
Purine Nucleotide Cycle ◽  
Acute Metabolic Acidosis ◽  
Ammonia Formation

To understand the mechanisms that initiate the increase in ammonia formation during acute acidosis in kidney [amino-15N]- and [amino-15N]glutamine were used as substrates in isolated perfused rat kidney experiments. Perfused kidneys from methionine sulfoximine-treated rats take up glutamine nitrogen at the rate of 1.50 +/- 0.08 mumol.g kidney-1.min-1 while forming ammonia at a rate of 0.65 +/- 0.09 mumol.g.kidney-1.min-1. Mass spectrometer analysis of the perfusate and urine reveals that ammonia is formed from the amide nitrogen of glutamine at the rate of 0.32 +/- 0.06 mumol.g kidney-1.min-1 and ammonia is formed from glutamate derived from glutamine at the rate of 0.21 +/- 0.04 mumol.g kidney-1.min-1. The balance of the ammonia formed is from unidentified endogenous sources. Addition of HCl to the perfusate to lower perfusate pH increases ammonia formation to 1.09 +/- 0.10 mumol.g kidney-1.min-1. The results exclude a role for the purine nucleotide cycle during acute acidosis and confirm that ammonia formation from glutamate derived from glutamine is via glutamate dehydrogenase. Lowering perfusate pH increases the rate of glutamine deamidation significantly by 0.33 +/- 0.06 mumol.g kidney-1.min-1 and increases the rate of ammonia formation via glutamate dehydrogenase insignificantly by only 0.08 +/- 0.04 mumol.g kidney-1.min-1, whereas ammonia formation from endogenous sources remains unchanged. The results demonstrate that regulation of glutamine deamidation is an important controlling step in ammonia formation during acute metabolic acidosis in kidney.

scholarly journals The purine nucleotide cycle and ammonia formation from glutamine by rat kidney slices

1983 ◽  
Vol 212 (3) ◽  
pp. 705-711 ◽  
Author(s):  
T Strzelecki ◽  
J Rogulski ◽  
S Angielski
Keyword(s):  
Adenine Nucleotides ◽  
Rat Kidney ◽  
Glucose Production ◽  
Amino Group ◽  
Purine Nucleotide ◽  
Normal Kidney ◽  
Kidney Slices ◽  
Total Ammonia ◽  
Purine Nucleotide Cycle ◽  
Ammonia Formation

To test the significance of the purine nucleotide cycle in renal ammoniagenesis, studies were conducted with rat kidney cortical slices using glutamate or glutamine labelled in the alpha-amino group with 15N. Glucose production by normal kidney slices with 2 mM-glutamine was equal to that with 3 mM-glutamate. With L-[15N]glutamate as sole substrate, one-third of the total ammonia produced by kidney slices was labelled, indicating significant deamination of glutamate or other amino acids from the cellular pool. Ammonia produced from the amino group of L-[alpha-15N]glutamine was 4-fold higher than from glutamate at similar glucose production rates. Glucose and ammonia formation from glutamine by kidney slices obtained from rats with chronic metabolic acidosis was found to be 70% higher than by normal kidney slices. The contribution of the amino group of glutamine to total ammonia production was similar in both types of kidneys. No 15N was found in the amino group of adenine nucleotides after incubation of kidney slices from normal or chronically acidotic rats with labelled glutamine. Addition of Pi, a strong inhibitor of AMP deaminase, had no effect on ammonia formation from glutamine. Likewise, fructose, which may induce a decrease in endogenous Pi, had no effect on ammonia formation. The data obtained suggest that the contribution of the purine nucleotide cycle to ammonia formation from glutamine in rat renal tissue is insignificant.

Effect of Decrease in Bicarbonate Concentration on Metabolism of the Isolated Perfused Rat Kidney

1979 ◽  
Vol 57 (1) ◽  
pp. 103-111 ◽  
Author(s):  
B. D. Ross ◽  
R. L. Tannen
Keyword(s):  
Metabolic Acidosis ◽  
Metabolic Regulation ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Rat Kidney ◽  
Ammonia Production ◽  
Perfusion Medium ◽  
Isolated Perfused Rat Kidney ◽  
Acid Urine ◽  
Perfused Rat Kidney ◽  
Acute Metabolic Acidosis

1. An isolated perfused rat kidney preparation which responds to acidification of the perfusion medium with the production of an acid urine and increased ammonia production was used to study the metabolic regulation of ammonia production from glutamine. 2. An inhibitor of gluconeogenesis at phosphoenolpyruvate carboxykinase(GTP), mercaptopicolinate, completely prevented the increase in ammoniagenesis, without preventing acidification of the urine. 3. Acidification of the perfusion medium from pH 7·4 to 7·0 reduced the renal concentrations of malate and 2-oxoglutarate. 4. Malate concentration was restored by inhibition of phosphoenolpyruvate carboxykinase(GTP), but 2-oxoglutarate content remained low. This indicates that accelerated gluconeogenesis in acute acidosis cannot be the explanation for the fall in 2-oxoglutarate concentration. 5. The fall in 2-oxoglutarate content is taken to indicate an important fall in tissue pH or in the redox ratio (NAD+/NADH) or both during acute metabolic acidosis. 6. From these studies with lowered bicarbonate two separate stimuli to ammoniagenesis in acute metabolic acidosis are postulated: urinary trapping of ammonia and increased disposal of glutamine carbon atoms via the pathway of glucose synthesis.

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