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T C Welbourne

Publications and source records attributed to T C Welbourne.

At least 55 records · Page 3Linked to original sources
PubMed1987

Interorgan glutamine flow in metabolic acidosis.

Acid-base homeostasis depends on glutamine flow from producer organs to those capable of generating bicarbonate. Glutamine oxidation, the prerequisite metabolic transformation, can be expressed by many sites; however, net base generation requires that glutamine flow be directed to a specific organ, the kidney. Normally, glutamine flows from the periphery to the splanchnic bed, providing a major fuel and supporting ureagenesis. Glutamine flow in chronic metabolic acidosis, on the other hand, is rerouted to the kidneys; asymmetrical distribution of NH+4 and HCO3- into the urine and renal vein subserves restoration of alkaline reserves. Clearly, glutamine flows in accordance with physiological demands, yet little is known of the regulatory mechanisms. As a model, chronic metabolic acidosis alters two aspects of this vital flow, its direction and magnitude. Characteristically the direction of flow is away from the splanchnic bed and into the kidneys associated with a marked fall in arterial glutamine concentration, restoring arterial level returns flow to the splanchnic bed sink. Thus glutamine homeostasis is sacrificed to impart direction to interorgan glutamine flow. Although multiple sites contribute to glutamine homeostasis, of great strategic importance is the potent hepatic glutaminase flux activated by portal venous NH+4 fed forward by gut metabolism; local hydrogen ion concentration modulates the effectiveness of this activator. Acute regulation of flow direction can be exerted by the lungs in determining the prevailing pCO2 and cellular acidity; respiratory compensation in chronic acidosis allows the expression of hepatic glutaminase, thereby suppressing arterial glutamine concentration. The enormous magnitude of glutamine flowing from muscle to the kidneys is supported by adaptive increases in glutamine synthetase and mitochondrial glutaminase, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Acid-Base Equilibrium↗
PubMed1986

Gamma glutamyltransferase ammonia production from glutamine: effect of physiological glutathione concentration.

The formation of ammonia from physiological glutamine concentration catalyzed by gamma glutamyltransferase was studied in the presence of physiological glutathione concentration. The apparent Km for ammonia formation from glutamine was 1.6 mM some 2 fold greater than the actual plasma concentration. In the presence of 30 microM glutathione neither the apparent Km or Vmax were changed. At supraphysiological glutathione concentration, 1mM, the apparent Km was increased while the Vmax decreased to one third. Hippurate the physiological modulator of the enzyme's glutaminase activity reduced the Km to 0.9 mM, the physiological range, and elevated the Vmax 2.7 fold.

Ammonia↗
PubMed1986

Fate of intraluminal glutamine in the perfused renal tubule.

To determine the fate of intraluminal glutamine and specifically the role of brush border gamma glutamyltransferase in its hydrolysis and reabsorption, proximal convoluted tubules of rabbits were isolated and perfused with an artificial ultrafiltrate containing 1 mM 14C-glutamine and 3H-PEG as a volume absorption marker. The tubules, average length 0.80 +/- 0.09 mm, were bathed in perfusate containing albumin, 6.5 percent but no glutamine. Aliquots of collectate and bathing media were monitored for total 14C counts while the distribution of radioactive 14C between glutamine and glutamate in the collectate was determined by separation on a Dowex X8 formate form ion-exchange column. After 3 ten minute control periods the perfusate was switched to one containing 1 mM AT-125 in addition to glutamine and after equilibration an additional 3 collections were obtained. Control period glutamine load averaged 16.1 +/- 2.4 pmole/min of which 35 percent was absorbed and 38 and 27 percent excreted as glutamine and glutamate respectively; of the absorbed glutamine 25 percent was metabolized. During AT-125 administration, glutamine delivery averaged 15.0 +/- 2.1 pmole/min of which 57 percent was absorbed; increased absorption occurred at the expence of intraluminal glutamate formation which fell to less than 10 percent. Thus luminal transport and gamma glutamyltransferase mediated hydrolysis appear to compete for available glutamine. Significantly, reducing intraluminal glutamine hydrolysis doubles the cellular metabolism of absorbed glutamine suggesting that extracellular conversion of glutamine to glutamate alters the metabolic fate of filtered glutamine.

Absorption↗
PubMed1986

Effect of metabolic acidosis on hindquarter glutamine and alanine release.

The relationship between hindquarter glutamine release and renal glutamine extraction was studied in rats undergoing chronic metabolic acidosis. Metabolic acidosis was induced by maintaining rats on NH4CL for four days; controls were pair-fed NH4HCO3. Metabolic acidosis increased renal glutamine extraction 8-fold and decreased arterial plasma glutamine concentration 40%. Hindquarter glutamine release rose 5-fold in acidosis due to an increased arteriovenous glutamine concentration difference and a significant rise in blood flow through the hindquarters. Both ammonia and glutamate extraction in the hindquarter increased in acidosis and could account for 25% of the glutamine nitrogen released. Alanine release by hindquarters, which exceeded glutamine release in the control animals, was greatly depressed in metabolic acidosis. The reduction in alanine nitrogen release during acidosis could account for 25% of the glutamine nitrogen released. Consequently, metabolic acidosis stimulates hindquarter glutamine release at the expense of alanine and is also supported by greater extraction of ammonia and glutamate. However, other N sources must supply nearly 50% of the glutamine nitrogen released based on balance studies.

Acidosis↗
PubMed1986

Renal acid-base metabolism after ischemia.

The response of the kidney to ischemia-induced cellular acidosis was followed over the immediate one hr post-ischemia reflow period. Clearance and extraction experiments as well as measurement of cortical intracellular pH (pHi) were performed on Inactin-anesthetized Sprague-Dawley rats. Arteriovenous concentration differences and para-aminohippurate extraction were obtained by cannulating the left renal vein. Base production was monitored as bicarbonate released into the renal vein and urine; net base production was related to the renal handling of glutamine and ammonia as well as to renal oxygen consumption and pHi. After a 15 min control period, the left renal artery was snared for one-half hr followed by release and four consecutive 15 min reflow periods. During the control period, cortical cell pHi measured by [14C]-5,5-Dimethyl-2,4-Oxazolidinedione distribution was 7.07 +/- 0.08, and Q-O2 was 14.1 +/- 2.2 micromoles/min; neither net glutamine utilization nor net bicarbonate generation occurred. After 30 min of ischemia, renal tissue pH fell to 6.6 +/- 0.15. However, within 45 min of reflow, cortical cell pH returned and exceeded the control value, 7.33 +/- 0.06 vs. 7.15 +/- 0.08. This increase in pHi was associated with a significant rise in cellular metabolic rate, Q-O2 increased to 20.3 +/- 6.4 micromoles/min. Corresponding with cellular alkalosis was a net production of bicarbonate and a net ammonia uptake and glutamine release; urinary acidification was abolished. These results are consistent with a nonexcretory renal metabolic base generating mechanism governing cellular acid base homeostasis following ischemia.

Acidosis, Renal Tubular↗
PubMed1986

Regulation of interorganal glutamine flow in metabolic acidosis.

Metabolic acidosis redirects interorgan glutamine flow from hepatic utilization to renal ammoniagenesis at the expense of ureagenesis. The roles of arterial glutamine load and organ glutaminase capacity in the regulation of glutamine balance across the gut, liver, and kidneys were studied in control and chronically acidotic rats. In control rats these organs combined to remove 733 nmol glutamine X min-1 X 100 g-1 in agreement with their respective glutaminase content, gut greater than liver greater than kidneys. In chronic metabolic acidosis renal glutamine extraction alone increased to 1,158 nmol X min-1 X 100 g-1 associated with an increased glutaminase capacity. However, the total glutamine deficit across these organs rose to only 1,043 nmol glutamine consumed X min-1 X 100 g-1 as a consequence of hepatic glutamine uptake reversing to net release. This reversal was not dependent on increased hepatic glutamine synthetase capacity, but rather appears to be dependent on the combined effect of reduced portal venous glutamine load and increased ammonia load. The reduction in portal glutamine load is, in turn, a consequence of renal glutamine extraction and reduced arterial glutamine concentration in metabolic acidosis as well as maintained gut glutamine extraction. Elevating arterial glutamine concentration in metabolic acidosis has no effect on renal uptake, but enhances splanchnic bed extraction with the restoration of ureagenesis. Thus the interorgan flow of glutamine and deposition of N into either urea or ammonia appears to be dependent on arterial glutamine concentration and hence glutamine availability in chronic metabolic acidosis in the rat.

Acid-Base Equilibrium↗
PubMed1986

Renal regulation of interorgan glutamine flow in metabolic acidosis.

The regulation of interorgan glutamine flow was studied in control and chronically metabolically acidotic rats. Net glutamine extraction or production across the kidneys, gut, liver, and hindquarters was determined in fasted anesthetized animals from organ blood flows and the arteriovenous glutamine concentration difference. In control animals glutamine flows from the hindquarters to the splanchnic bed. In chronic acidosis glutamine production by the hindquarters rose threefold and was redirected to the kidneys; splanchnic bed glutamine uptake was eliminated. Associated with this was a 39% fall and a 62% rise in arterial glutamine and ammonia concentrations, respectively. Removing the kidneys from the circulation returned arterial glutamine and ammonia concentrations to control nonacidotic levels within 30 min. Net glutamine production by the hindquarters decreased, whereas splanchnic bed glutamine extraction increased. Hindquarter glutamine production appears to be modulated by renal venous ammonia; splanchnic bed glutamine extraction is load dependent, reflecting the influence of renal glutamine consumption on the steady-state arterial levels. Thus the removal of the kidneys returns interorgan glutamine flow to that observed in nonacidotic animals consistent with a major role of the kidneys in regulating glutamine flow and nitrogen metabolism in chronic metabolic acidosis.

Acidosis↗
PubMed1986

Hepatic glutaminase flux regulation of glutamine homeostasis. Studies in vivo.

The role of hepatic glutaminase flux in regulating plasma glutamine homeostasis was studied in the intact rat. Interorgan glutamine flow during chronic metabolic acidosis was away from the splanchnic bed and to the kidneys. Hindquarter and hepatic glutamine release were the major sources of glutamine removed by the kidneys. Interorgan glutamate flow was from the liver to the hindquarters and kidneys. Chronic metabolic acidosis reduced arterial glutamine concentration 30%. Acute respiratory acidosis (pH 7.12 +/- 0.02) returned arterial glutamine concentration to normal values, increasing and decreasing hepatic glutamine and glutamate release respectively; renal and gut glutamine removal rates were not decreased. Hepatic unidirectional glutamine utilization measured isotopically was decreased 51% by acute acidosis; unidirectional glutamine production was unchanged. The results are consistent with the proposed role of ammonia-activated hepatic glutaminase in the regulation of glutamine homeostasis during acute acidosis.

Acid-Base Equilibrium↗
PubMed1986

Hepatic enzymes of glutamine and ureagenesis in metabolic acidosis.

The activities of key glutamine and urea cycle enzymes were assayed in liver homogenates from control and chronically acidotic rats and compared with citrulline and urea productions by isolated mitochondria and intact liver slices, respectively. Glutamine-dependent urea and citrulline synthesis were increased significantly in isolated mitochondria and in liver slices; the activities of carbamoyl phosphate synthetase and arginase were unchanged and increased, respectively. Glutamine was not a precursor in the carbamoyl phosphate synthetase system, suggesting that the glutamine effect is an indirect one and that glutamine requires prior hydrolysis. Increased mitochondrial citrulline synthesis was associated with enhanced oxygen consumption, suggesting glutamine acts both as a nitrogen and fuel source. Hepatic phosphate-dependent glutaminase was elevated by chronic acidosis. The results indicate that the acidosis-induced reduction in ureagenesis and reversal from glutamine uptake to release observed in vivo are not reflections of corresponding changes in the hepatic enzyme content. Rather, when available, glutamine readily supports ureagenesis, suggesting a close coupling of hepatic glutaminase flux with citrulline synthesis.

Acidosis↗
PubMed1985

Renal sodium reabsorption, oxygen consumption, and gamma-glutamyltransferase excretion in the postischemic rat kidney.

The effect of 30 min of renal artery occlusion on renal sodium reabsorption, oxygen consumption, and brush border integrity was studied in the immediate 1-h reflow period. Rats were studied using clearance and renal extraction measurements. Glomerular filtration rate and renal plasma flow were measured over four consecutive 15-min periods; renal venous samples were drawn via an indwelling catheter. Renal oxygen consumption (QO2) was calculated from renal blood flow, corrected for urine flow, and from blood oxygen content measured with a fuel cell analyzer. Brush border integrity was assessed by the excretion of the brush border marker enzyme gamma-glutamyltransferase as well as by morphologic observation. Ischemia induced a 10-fold rise in fractional sodium excretion in the initial 0- to 15-min period, rose to 20-fold during the subsequent two periods, 15-45 min, and then returned to the initial reflow period level. The progressive deterioration of renal function with reflow could not be attributed to an increase in the filtered Na+ load. Rather, Na+ reabsorption appeared to be related to the presence of intact brush borders at 0-15 min, their removal from the luminal surface between 15 and 45 min, and their return at 60 min of reflow. Renal QO2 was coupled to Na+ reabsorption in the initial 15-min and final 45- to 60-min reflow periods. However QO2 was significantly increased over the control level at 30-45 min of reflow. The results point to a significant role of brush border uptake in the development of functional impairment following renal ischemia and suggest that the associated rise in renal O2 consumption may be coupled to the reparation of this organelle.

Absorption↗
PubMed1984

Glutamine homeostasis: role of pCO2 in regulating arterial glutamine in metabolic acidosis.

In chronic metabolic acidosis arterial plasma glutamine concentration is reduced 46%. This effect is attributable to a reduction in arterial pCO2 rather than acidemia since elevating pCO2 returns glutamine toward the control level. The extracellular glutamine precursors ammonia and glutamate exhibit a reciprocal decline with the elevation in pCO2. This reduction in metabolic acidosis appears to play an important role in sparing glutamine for renal base generation.

Acidosis↗
PubMed1983

Gamma-glutamyltransferase release by the post ischemic kidney: multiple forms and cellular pH.

The release of gamma-glutamyltransferase from renal tubule cells was studied in situ following 30 minutes of ischemia. The ischemic kidney enzyme level fell 33 percent after 15 minutes of reflow of which only 1.2 percent was recovered in the urine; none was released into the renal vein. At this time the overwhelming majority of the enzyme appears bound to membranes in both the kidney and the urine. In the subsequent 15 minutes renal levels continue to decline while urinary excretion accounts for 5 percent of that disappearing from the kidney. Interestingly the form of the enzyme present in kidney and urine shifts to a soluble form coinciding with cellular alkalosis, urinary alkalinization and a rise in ATP levels. Alkalinization of renal homogenates result in a 2-fold increase in the soluble enzyme form. The results are consonant with the immediate loss of brush border enzyme via uptake into the cell or release into the urine with the former pathway predominating; subsequent appearance of the soluble enzyme appears to reflect intracellular alkaline proteinase activity and exocytosis. The form in which the enzyme is excreted may provide a useful clinical index: membranous reflecting cellular necrosis and soluble reflecting cellular recovery.

Acyltransferases↗
PubMed1982

Acetazolamide inhibition of renal gamma-glutamyl transpeptidase.

The effect of acetazolamide (AZ) on renal gamma-glutamyl transpeptidase (EC 2.3.2.2) activity (gamma-GT) was studied with the purified enzyme, subcellular fractions, and in the isolated functioning kidney. Activity of gamma-GT was assessed using either one of two gamma-glutamyl donors, gamma-glutamyl-p-nitroanilide (gamma GpNA) or glutamine, and either the gamma-glutamyl acceptor glycylglycine (Gly-Gly) or methionine (Met). With the microsomal enzyme and beta-GpNA, AZ was shown to inhibit p-nitroaniline (p-NA) formation; however, gamma-GpNA Km remained unchanged (1.8 mM), while the Vmax was reduced significantly, 333 vs 200 mumoles . min-1 . mg-1. Adding Gly-Gly removed AZ inhibition, while AZ elevated the apparent Km from Gly-Gly from 16 to 48; AZ inhibition of gamma-GT activity resulted in a decrease in gamma-glutamyl-Gly-Gly formation consistent with interaction at the gamma-glutamyl acceptor site. With glutamine as the beta-glutamyl donor, AZ reduced NH3 and apparent gamma-glutamylmethionine formation in the purified enzyme in agreement with inhibition at the acceptor site. In the functioning kidney, perfused with 10(-3)M L- or D-glutamine, AZ (10(-3)M) markedly reduced NH3 formation and increased glutamine excretion, results consistent with AZ inhibition of the in situ gamma-GT.

Acetazolamide↗
PubMed1982

2,4-dinitrophenol stimulation of renal ammoniagenesis.

The role of mitochondrial swelling in renal ammoniagenesis was studied by the administration of 10 mg.kg-1 2,4-dinitrophenol, in vivo, to normal and chronically acidotic rats. 2,4-Dinitrophenol increased ammonia excretion in in vivo and in vitro production from glutamine by renal cortical slices and isolated kidneys perfused with 1 mM L-glutamine. Ammonia production per glutamine molecule utilized rose towards 2, consistent with activation of the mitochondrial glutaminase-glutamate dehydrogenase pathway in 2,4-dinitrophenol-treated and acidotic rats. The rank order of 2,4-dinitrophenol stimulation of ammonia formation in vivo and in vitro was normal less than normal + 2,4-dinitrophenol less than acidotic less than acidotic + 2,4-dinitrophenol. 2,4-Dinitrophenol administration appears to enlarge the in situ proximal tubule mitochondrial population and to increase the number undergoing degradation, suggesting that mitochondrial alterations correlate with ammoniagenesis in vivo.

2,4-Dinitrophenol↗
PubMed1982

Function of renal gamma-glutamyltransferase: significance of glutathione and glutamine interactions.

The membrane bound gamma-glutamyltransferase (gamma-GT) is capable of utilizing both glutathione, GSH, and glutamine, gln, as the natural gamma-glutamyl donor. The enzyme is oriented in the membrane to react with extracellular substrates and is present on both the brush border and peritubular capillaries. The reaction catalyzed by gamma-GT is critically dependent upon the ratio of gamma-glutamyl donor/gamma-GT which under near physiological conditions results in the formation of gamma-glutamyl peptide and glu on the blood and urine side respectively; this effectively establishes an osmotic gradient which could contribute to transepithelial and transcapillary water fluxes. Interestingly, utilization of extrarenal gamma-glutamyl substrates are quantitatively more significant in the microvascular than brush border location. Delivery of gln to these enzymes sites is some 20 times greater than GSH. Gln utilization unlike GSH is limited by the reaction with the gamma-glutamyl donor site; thus reactivity is greatly enhanced by the maleate like activator hippurate which may account for the acidosis-induced adaptation in ammonia formation from gln by this enzyme. Coupled to a role in ammoniagenesis the brush border enzyme appears to play a role in the reabsorption of filtered gln. The hydrolysis of filtered GSH as well as its utilization in transfer reactions involved in amino acid reabsorption at the brush border may reflect the role of the enzyme in eliminating osmotically active solutes from the urine and thereby facilitating water fluxes. However the role of gln as a gamma-glutamyl donor relative to GSH will depend upon the quantitative significance of tubular GSH synthesis and secretion.

Animals↗