Activities Of Urea Cycle Enzymes Research Articles

Ureagenesis of Asian seabass (Lates calcarifer) under ammonia stress and overcrowding

Ammonia, an excretory substance resulting from the catabolism of proteins and amino acids, is toxic to fish. Increasing ammonia, due to overcrowding fish in a tank, results in dropping production efficiency. This study aims to investigate the adaptation of Asian sea bass to increased ammonia content in fish tanks with different populations. Five groups were tested in 250-l plastic tanks: group A: 15 fish (6 g/L), group B: 20 fish (8 g/L), group C: 25 fish (10 g/L), group D: 30 fish (12 g/L), group E: 35 fish (14 g/L). The ammonia level in the tanks was measured daily. After 60 days, the activity of some enzymes and biochemical parameters related to ammonia was measured in muscle, liver, and blood. The results were statistically analyzed. After hypoxia, in the overcrowded group, the level of liver glycogen and blood glucose fell; the blood lactate and muscle adenosine monophosphate deaminase (AMPD) activity increased; the activity of muscle glutamate dehydrogenase (GDH) increased; the activity of glutamate dehydrogenase, glutaminase, arginase, and aspartate transferase (AST) of the liver increased. Due to the increased activity of urea cycle enzymes in the liver, urea was detected in the blood. One of the strategies that Asian sea bass adopts in dealing with a high level of ammonia is to activate the urea cycle in the liver and turn ammonia into urea to control it. This way is a type of adaptation to ammonia stress that was not identified in this species before.

A curious case of confusion

A curious case of confusion

Ammonia Scavenging Prevents Progression of Fibrosis in Experimental Nonalcoholic Fatty Liver Disease.

In nonalcoholic fatty liver disease (NAFLD), fibrosis is the most important factor contributing to NAFLD-associated morbidity and mortality. Prevention of progression and reduction in fibrosis are the main aims of treatment. Even in early stages of NAFLD, hepatic and systemic hyperammonemia is evident. This is due to reduced urea synthesis; and as ammonia is known to activate hepatic stellate cells, we hypothesized that ammonia may be involved in the progression of fibrosis in NAFLD. In a high-fat, high-cholesterol diet-induced rodent model of NAFLD, we observed a progressive stepwise reduction in the expression and activity of urea cycle enzymes resulting in hyperammonemia, evidence of hepatic stellate cell activation, and progressive fibrosis. In primary, cultured hepatocytes and precision-cut liver slices we demonstrated increased gene expression of profibrogenic markers after lipid and/or ammonia exposure. Lowering of ammonia with the ammonia scavenger ornithine phenylacetate prevented hepatocyte cell death and significantly reduced the development of fibrosis both in vitro in the liver slices and in vivo in a rodent model. The prevention of fibrosis in the rodent model was associated with restoration of urea cycle enzyme activity and function, reduced hepatic ammonia, and markers of inflammation. The results of this study suggest that hepatic steatosis results in hyperammonemia, which is associated with progression of hepatic fibrosis. Reduction of ammonia levels prevented progression of fibrosis, providing a potential treatment for NAFLD.

The Food Energy/Protein Ratio Regulates the Rat Urea Cycle but Not Total Nitrogen Losses.

Nitrogen balance studies have shown that a portion of the N ingested but not excreted is not accounted for. We compared several diets (standard, high-fat, high-protein, and self-selected cafeteria) to determine how diet-dependent energy sources affect nitrogen handling, i.e., the liver urea cycle. Diet components and rat homogenates were used for nitrogen, lipid, and energy analyses. Plasma urea and individual amino acids, as well as liver urea cycle enzyme activities, were determined. Despite ample differences in N intake, circulating amino acids remained practically unchanged in contrast to marked changes in plasma urea. The finding of significant correlations between circulating urea and arginine-succinate synthase and lyase activities supported their regulatory role of urea synthesis, the main N excretion pathway. The cycle operation also correlated with the food protein/energy ratio, in contraposition to total nitrogen losses and estimated balance essentially independent of dietary energy load. The different regulation mechanisms observed have potentially important nutritional consequences, hinting at nitrogen disposal mechanisms able to eliminate excess nitrogen under conditions of high availability of both energy and proteins. Their operation reduces urea synthesis to allow for a safe (albeit unknown) mechanism of N/energy excess accommodation.

Distributional shift of urea production site from the extraembryonic yolk sac membrane to the embryonic liver during the development of cloudy catshark (Scyliorhinus torazame)

Urea is an essential osmolyte for marine cartilaginous fishes. Adult elasmobranchs and holocephalans are known to actively produce urea in the liver, muscle and other extrahepatic organs; however, osmoregulatory mechanisms in the developing cartilaginous fish embryo with an undeveloped urea-producing organ are poorly understood. We recently described the contribution of extraembryonic yolk sac membranes (YSM) to embryonic urea synthesis during the early developmental period of the oviparous holocephalan elephant fish (Callorhinchus milii). In the present study, to test whether urea production in the YSM is a general phenomenon among oviparous Chondrichthyes, we investigated gene expression and activities of ornithine urea cycle (OUC) enzymes together with urea concentrations in embryos of the elasmobranch cloudy catshark (Scyliorhinus torazame). The intracapsular fluid, in which the catshark embryo develops, had a similar osmolality to seawater, and embryos maintained a high concentration of urea at levels similar to that of adult plasma throughout development. Relative mRNA expressions and activities of catshark OUC enzymes were significantly higher in YSM than in embryos until stage 32. Concomitant with the development of the embryonic liver, the expression levels and activities of OUC enzymes were markedly increased in the embryo from stage 33, while those of the YSM decreased from stage 32. The present study provides further evidence that the YSM contributes to embryonic urea homeostasis until the liver and other extrahepatic organs become fully functional, and that urea-producing tissue shifts from the YSM to the embryonic liver in the late developmental period of oviparous marine cartilaginous fishes.

Influence of feeding increasing levels of dry or modified wet corn distillers’ grains plus solubles in whole corn grain-based finishing diets on hepatic and renal mass, and glutathione peroxidase and urea cycle enzyme activities in finishing cattle

Salim, H., Wood, K. M., Cant, J. P. and Swanson, K. C. 2015. Influence of feeding increasing levels of dry or modified wet corn distillers’ grains plus solubles in whole corn grain-based finishing diets on hepatic and renal mass, and glutathione peroxidase and urea cycle enzyme activities in finishing cattle. Can. J. Anim. Sci. 95: 407–415. Forty-two cross-bred steers (BW=357±5.8 kg) fed whole corn grain-based finishing diets were used in a completely randomized block (60, 120, or 180 d on feed) design (2×3 factorial arrangement of treatments plus control) to determine the effect of inclusion level [0 (control), 16.7, 33.3, and 50% of diet DM) and form (dry (DDGS) or modified wet (MWDGS)] of distillers’ grains plus solubles (DGS) on hepatic and renal glutathione peroxidase (GPx) and hepatic urea cycle enzyme activities. Kidney weight (g kg−1of BW) increased linearly (P=0.004) with increasing inclusion levels of DGS. There were no effects (P≥0.11) of dietary treatment on hepatic and renal GPx activity (U g−1, U mg−1of protein, and kU liver−1). Hepatic carbamoyl phosphate synthetase activity (kU liver−1and U kg−1of BW) tended to linearly increase (P=0.09 and P=0.10, respectively) with increasing inclusion level of DGS. Hepatic ornithine transcarbamoylase and argininosuccinate synthetase activity (kU liver−1and U kg−1of BW) increased linearly (P≤0.05) with increasing inclusion levels of DGS. These data indicate that steers adapt to feeding up to 50% DGS by increasing kidney mass and activity of urea cycle enzymes in liver to allow for clearance of excess nitrogen. Also, hepatic and renal GPx activity, as an indicator of Se status, is not affected when typical finishing diets are fed.

Methane digester effluent from swine excreta as a nutrient and water source for growing and finishing swine.

The experimental objective was to evaluate swine methane digester effluent (SMDE) as a water and nutrient source for swine. The mesophilic methane digester was loaded daily with manure from finishing swine fed a corn-soybean meal diet. Dry diet was mixed with SMDE (3.7% DM) and fed twice daily in troughs. Tap water was provided and consumption measured. Barrows were group fed (3 pigs/pen) and adapted to SMDE by increasing SMDE for 7 d, with the full amount fed from d 8 to the end of the feeding phase (d 21, 14, 23, or 37 for Exp. 1 to 4, respectively). Blood samples were collected on d 0, 10, 21, and 31 to determine plasma concentrations of glucose and plasma urea N (PUN). Barrows were placed in individual metabolism cages for a 5-d acclimation and a 5-d fecal and urine collection to determine apparent N and energy utilization. For Exp. 1, 18 pigs averaging 75 kg BW were allotted to diets with 0, 48.6, or 63.7% SMDE, as-fed basis. For Exp. 2 and 3, 12 pigs/experiment averaged 117 and 70 kg, respectively, and were allotted to diets with 0 or 63.7% SMDE, as-fed basis. At the end of Exp. 2 and 3, pigs were sacrificed and liver samples were collected to determine urea cycle enzyme activity, and loin was saved for taste panel evaluation. For Exp. 4, pigs averaged 40 kg and were allotted to diets with 0 or 57.5% SMDE, as-fed basis. The ADFI, ADG, and G:F of finishing swine (Exp. 1 to 3) were not reduced by feeding diets containing 63.7% SMDE (as-fed basis), whereas ADG and G:F of growing swine (Exp. 4) were reduced (P < 0.01) by feeding a diet containing 57.5% SMDE. Pigs fed diets containing SMDE consumed 31 to 56% less (P < 0.05) water and had greater (P < 0.01) PUN concentrations than pigs fed control diets. Pigs fed diets containing SMDE excreted more (g, P < 0.05) fecal N and absorbed and retained less N (%; P < 0.01) and energy (DE and ME) than pigs fed control diets. Treatment had no effect on urea cycle enzyme activity. In conclusion, finishing swine adapted to diets containing 63.7% SMDE (as-fed basis) based on growth performance, whereas growing swine did not adapt to a diet containing 57.5% SMDE because of the large content of nonprotein N in SMDE. Recycling SMDE to swine greatly reduced fresh water consumption, whereas the protein and energy values of SMDE were approximately 0 for swine. Therefore, SMDE is more appropriately recycled as a source of water and N for ruminant nutrition or crop production.

Effect of the Quality of Dietary Amino Acids Composition on the Urea Synthesis in Rats

We have shown that urinary urea excretion increased in rats given a lower quality protein. The purpose of present study was to determine whether the composition of dietary amino acids affects urea synthesis. Experiments were done on three groups of rats given diets containing a 10% gluten amino acid mix diet or 10% casein amino acid mix diet or 10% whole egg protein amino acids mix diet for 10 d. The urinary excretion of urea, the liver concentration of N-acetylglutamate, and the liver concentration of free serine, glutamic acids and alanine were greater in the group given the amino acid mix diet of lower quality. The fractional and absolute rates of protein synthesis in tissues declined with a decrease in quality of dietary amino acids. The hepatic concentration of ornithine and the activities of hepatic urea-cycle enzymes were not related to the urea excretion. These results suggest that the increased concentrations of amino acids and N-acetylglutamate seen in the liver of rats given the amino acid mix diets of lower quality are likely among the factors stimulating urea synthesis. The protein synthesis in tissues is at least partly related to hepatic concentrations of amino acids. The composition of dietary amino acids is likely to be one of the factors regulating urea synthesis when the quality of dietary protein is manipulated.

Effects of crowding on ornithine–urea cycle enzyme mRNA expression and activity in gulf toadfish (Opsanus beta)

The gulf toadfish (Opsanus beta) is a facultatively ureotelic fish that excretes primarily urea under conditions of crowding or confinement. To examine the relationship between ammonia production, urea production and the ornithine-urea cycle (O-UC) enzyme activity and mRNA expression, we subjected toadfish to two-day and seven-day crowding regimes. Plasma cortisol levels were measured and liver tissue was assayed for ammonia and urea concentrations. Liver glutamine synthetase (GS), carbamoyl phosphate synthetase III (CPS), ornithine carbamoyl transferase (OCT) and arginase (ARG) activities were also measured. Quantitative PCR was utilized to determine liver GS, CPS, OCT, ARG, argininosuccinate synthetase (ASS) and argininosuccinate lyase (ASL) mRNA expression. Hepatic ammonia concentrations decreased with increased duration of crowding whereas liver urea and circulating cortisol levels increased. An elevation in enzyme activity with increased duration of crowding was observed for all four O-UC enzymes examined. By contrast, mRNA expression was variable for the O-UC enzymes and only CPS and ASS had mRNA expression levels that were elevated in crowded fish. These results suggest that the activities of O-UC enzymes are better predictors for urea production than O-UC enzyme mRNA expression levels.

Lysine 88 Acetylation Negatively Regulates Ornithine Carbamoyltransferase Activity in Response to Nutrient Signals

Ornithine carbamoyltransferase (OTC) is a key enzyme in the urea cycle to detoxify ammonium produced from amino acid catabolism. OTC deficiency is an X-linked genetic disorder ranging from fatal in newborns to hyperammonemia and anorexia in adults. Through affinity purification of acetylated peptides and mass spectrometry, we identified that OTC is acetylated on lysine residues, including Lys88, which is also mutated in OTC-deficient patients. OTC acetylation was confirmed to occur under physiological conditions. Biochemical characterizations revealed that OTC Lys88 acetylation decreases the affinity for carbamoyl phosphate, one of the two OTC substrates, and the maximum velocity, whereas the K(m) for ornithine, the other OTC substrate, is not affected. Furthermore, Lys88 acetylation is regulated by both extracellular glucose and amino acid availability, indicating that OTC activity may be regulated by cellular metabolic status. Our results provide an example of the novel mechanism of regulating metabolic enzyme activity through protein acetylation.

Changes of activity and mRNA expression of urea cycle enzymes in the liver of developing Holstein calves1

Urea is an important reutilizable nitrogen source for the ruminant and is mainly synthesized through the urea cycle in the liver. The cycle is undertaken by 5 enzymes: carbamoyl phosphate synthetase (CPS), ornithine transcarbamoylase (OTC), arginino-succinate synthetase (AS), argininosuccinate lyase (AL), and arginase. The purpose of this study was to investigate changes in the activity of the enzymes and mRNA expression, given that previous observations have indicated an increase in plasma urea concentrations with age in Holstein calves. First, plasma concentrations of metabolites and hormones were determined in calves at 1, 3, 8, 13, and 19 wk of age (n = 4, weaned at 6 wk of age). The plasma concentration of urea drastically increased after weaning (P < 0.001). The plasma concentration of glucose was lowest at 8 wk. The plasma concentration of IGF-I gradually increased with age, although those of NEFA, glucagon, and cortisol decreased (P < 0.001). Concentrations of triglyceride, alpha-amino nitrogen, growth hormone, and insulin did not change significantly with age of the calf. Next, using the liver tissues taken from calves at 2, 13, and 19 wk of age (n = 4 to 6 at each time point, weaned at 6 wk of age), we measured the activity and mRNA expression of the enzymes by biochemical methods and quantitative reverse transcription-PCR, respectively. The activities of CPS (P < 0.001), OTC (P = 0.001), and AS (P = 0.015) increased with age, whereas AL (P = 0.003) decreased. Although mRNA expression was decreased with age for AL (P = 0.002) and arginase (P = 0.007), no significant change was observed for CPS, OTC, or AS mRNA expression. We conclude that the increased urea production in the liver may be explained not only by an increase in the activities of the urea cycle enzymes, but also by increased ammonia production by rumen fermentation and gluconeogenesis from amino acids around weaning time.

Urea cycle enzymes through the development of pacu (Piaractus mesopotamicus): the role of ornithine carbamoyl transferase

The present work reports the activities of urea cycle enzymes during the ontogenic development of the teleost pacu (Piaractus mesopotamicus). Urea cycle enzymes from the kidney and liver of adult fish were compared with those from the fish's embryonic phases. Samples were evaluated over all phases of embryonic development, the larval period and alevin. Ammonia and urea concentrations were determined during embryogenesis and in the plasma of adult fish. Except for carbamoyl phosphate synthetase-III (CPS-III), all enzymes of the urea cycle were expressed in the larvae and alevins as well as in the liver and kidney of adult fish. In spite of the low level of activity of the ornithine urea cycle (OUC) enzymes compared to those in mammals, and the low levels of tissue urea concentration compared to ammonia, the ureogenesis was evaluated in pacu. Ammonia seems to be the main nitrogenous waste during embryonic development. In this phase glutamine synthetase (GS) may play a role in ammonia detoxification, and the OUC enzymes can be individually involved in functions other than urea production. The presence of ornithine carbamoyl transferase (OCT) in all developmental phases of pacu and in the adult liver and kidney suggests that this enzyme is performing different metabolic pathways. OCT in the kidney, wherein the activity is less than in the liver, should work in the biosynthesis of polyamines and control the arginine plasma concentration given that renal arginase and argininosuccinate synthetase-argininosuccinate lyase are more active than from the liver. We suppose that OCT during the embryogenesis is a control step regulating the cellular concentration of ornithine for polyamines synthesis.

The Dogfish Shark (Squalus acanthias) Increases both Hepatic and Extrahepatic Ornithine Urea Cycle Enzyme Activities for Nitrogen Conservation after Feeding

Urea not only is utilized as a major osmolyte in marine elasmobranchs but also constitutes their main nitrogenous waste. This study investigated the effect of feeding, and thus elevated nitrogen intake, on nitrogen metabolism in the Pacific spiny dogfish Squalus acanthias. We determined the activities of ornithine urea cycle (O-UC) and related enzymes in liver and nonhepatic tissues. Carbamoyl phosphate synthetase III (the rate-limiting enzyme of the O-UC) activity in muscle is high compared with liver, and the activities in both tissues increased after feeding. The contribution of muscle to urea synthesis in the dogfish body appears to be much larger than that of liver when body mass is considered. Furthermore, enhanced activities of the O-UC and related enzymes (glutamine synthetase, ornithine transcarbamoylase, arginase) were seen after feeding in both liver and muscle and were accompanied by delayed increases in plasma urea, trimethylamine oxide, total free amino acids, alanine, and chloride concentrations, as well as in total osmolality. The O-UC and related enzymes also occurred in the intestine but showed little change after feeding. Feeding did not change the rate of urea excretion, indicating strong N retention after feeding. Ammonia excretion, which constituted only a small percentage of total N excretion, was raised in fed fish, while plasma ammonia did not change, suggesting that excess ammonia in plasma is quickly ushered into synthesis of urea or protein. In conclusion, we suggest that N conservation is a high priority in this elasmobranch and that feeding promotes ureogenesis and growth. Furthermore, exogenous nitrogen from food is converted into urea not only by the liver but also by the muscle and to a small extent by the intestine.

Gene expression and activity of urea cycle enzymes of rat hepatocytes cold stored up to 120 h in University of Wisconsin solution

Urea cycle (UC) is the main pathway of ammonium removal. A deficiency in any of the five classical enzymes of the pathway causes a urea cycle disorder. Hepatocellular transplantation is one of the techniques applicable to treat this disorder. In the present work, we investigated the activities and the relative expression levels of two of the UC enzymes: Carbamyl phosphate synthetase I (CPSI) and ornithine transcarbamylase (OTC), in isolated hepatocytes preserved up to 120 h in University of Wisconsin (UW) solution at 0 °C, and during the rewarming of these suspensions. During preservation, CPSI showed differences in mRNA levels respect to time 0, while ornithine transcarbamylase remained unchanged. At the end of the rewarming, CPSI showed values of enzymatic activity and relative mRNA level comparable with the control, meanwhile, there was an increment in OTC activity. In line with these results, we found that hepatocytes cold preserved up to 120 h in UW solution maintained their ability to remove an ammonium load comparable to freshly isolated hepatocytes. These data indicated that our preservation conditions up to 120 h in UW solution followed by rewarming, preserves UC enzymes at levels similar to freshly isolated hepatocytes, allowing the use of these cells in bioartificial liver devices or hepatocellular transplantation.

Effect of adding dietary L-lysine, L-threonine and L-methionine to a low gluten diet on urea synthesis in rats

We have shown that urinary urea excretion increased in rats fed a low quality protein. The purpose of present study was to determine whether an addition of dietary limiting amino acids affected urea synthesis in rats fed a low gluten diet. Experiments were done on three groups of rats given diets containing 10% gluten, 10% gluten +0.5% L-lysine or 10% gluten+0.5% L-lysine, 0.2% L-threonine and 0.2% L-methionine for 10 d. The urinary excretion of urea, and the liver concentrations of serine and ornithine decreased with the addition of dietary L-lysine, L-threonine and L-methionine. The fractional and absolute rates of protein synthesis in tissues increased with the treatment of limiting amino acids. The activities of hepatic urea-cycle enzymes was not related to the urea excretion. These results suggest that the addition of limiting amino acids for the low gluten diet controls the protein synthesis in tissues and hepatic ornithine and decline urea synthesis.

Lamprey parasitism of sharks and teleosts: high capacity urea excretion in an extant vertebrate relic

We observed 10 sea lampreys ( Petromyzon marinus) parasitizing basking sharks ( Cetorhinus maximus), the world's second largest fish, in the Bay of Fundy. Due to the high concentrations of urea in the blood and tissues of ureosmotic elasmobranchs, we hypothesized that sea lampreys would have mechanisms to eliminate co-ingested urea while feeding on basking sharks. Post-removal urea excretion rates ( J Urea) in two lampreys, removed from separate sharks by divers, were initially 450 (≈9000 μmol N kg −1 h −1) and 75 times (≈1500 μmol N kg −1 h −1) greater than basal (non-feeding) rates (≈20 μmol N kg −1 h −1). In contrast, J Urea increased by 15-fold after parasitic lampreys were removed from non-ureosmotic rainbow trout ( Oncorhynchus mykiss). Since activities of the ornithine urea cycle (OUC) enzymes, carbamoyl phosphate synthetase III (CPSase III) and ornithine carbamoyl transferase (OCT) were relatively low in liver and below detection in intestine and muscle, it is unlikely that the excreted urea arose from de novo urea synthesis. Measurements of arginase activity suggested that hydrolysis of dietary arginine made a minor contribution to J Urea.. Post-feeding ammonia excretion rates ( J Amm) were 15- to 25-fold greater than basal rates in lampreys removed from both basking sharks and rainbow trout, suggesting that parasitic lampreys have a high capacity to deaminate amino acids. We conclude that the sea lamprey's ability to penetrate the dermal denticle armor of sharks, to rapidly excrete large volumes of urea and a high capacity to deaminate amino acids, represent adaptations that have contributed to the evolutionary success of these phylogenetically ancient vertebrates.

Role of N-acetylglutamate concentration and ornithine transport into mitochondria in urea synthesis of rats given proteins of different quality.

The purpose of this study was to find whether the concentration of N-acetylglutamate and ornithine transport into mitochondria would regulate urea synthesis when the dietary protein quality was manipulated. Experiments were done on three groups of rats given diets containing 10 g of gluten, 10 g of casein, or 10 g of whole egg protein/100 g for 10 days. The plasma concentration and urinary excretion of urea, the liver concentration and synthesis of N-acetylglutamate, the liver concentrations of glutamate and lysine, and the liver ornithine transport into mitochondria increased with the decrease in quality of dietary protein. A reverse correlation was observed between the activities of urea cycle enzymes, the plasma concentration of arginine, and urinary excretion of urea under these conditions. N-Acetylglutamate concentration and ornithine transport into mitochondria in the liver were closely correlated with the excretion of urea. These results suggest that greater N-acetylglutamate concentration and ornithine transport into isolated mitochondria in the liver of rats, given the lower quality of protein, stimulate urea synthesis and that the concentrations of glutamate and lysine in the liver are at least partly related to the hepatic N-acetylglutamate synthesis and ornithine transport, respectively.

Effect of alkalinity (pH 10) on ureogenesis in the air-breathing walking catfish, Clarias batrachus

Exposure of fish to alkaline conditions inhibits the rate of ammonia excretion, leading to ammonia accumulation and toxicity. The purpose of this study was to determine the role of ureogenesis via the urea cycle, to avoid the accumulation of ammonia to a toxic level during chronic exposure to alkaline conditions, for the air-breathing walking catfish, Clarias batrachus, where a full complement of urea cycle enzyme activity has been documented. The walking catfish can survive in water with a pH up to 10. At a pH of 10 the ammonia excretion rate by the walking catfish decreased by approximately 75% within 6 h. Although there was a gradual improvement of ammonia excretion rate by the alkaline-exposed fish, the rate remained 50% lower, even after 7 days. This decrease of ammonia excretion was accompanied by a significant accumulation of ammonia in plasma and body tissues (except in the brain). Urea-N excretion for alkaline-exposed fish increased 2.5-fold within the first day, which was maintained until day 3 and was then followed by a slight decrease to maintain a 2-fold increase in the urea-N excretion rate, even after 7 days. There was also a higher accumulation of urea in plasma and other body tissues (liver, kidney, muscle and brain). The activity of glutamine synthetase and three enzymes operating in the urea cycle (carbamyl phosphate synthetase, argininosuccinate synthetase, argininosuccinate lyase) increased significantly in hepatic and extra-hepatic tissue, such as the kidney and muscle in C. batrachus, during exposure to alkaline water. A significant increase in plasma lactate concentration noticed during alkaline exposure possibly helped in the maintenance of the acid–base balance. It is apparent that the stimulation of ureogenesis via the induced urea cycle is one of the major physiological strategies adopted by the walking catfish ( C. batrachus) during chronic exposure to alkaline water, to avoid the in vivo accumulation of ammonia to a toxic level in body tissues and for the maintenance of pH homeostasis.

Somatotropin-Induced Amino Acid Conservation in Pigs Involves Differential Regulation of Liver and Gut Urea Cycle Enzyme Activity

Somatotropin (ST) treatment promotes animal growth and allows for the conservation of amino acids by increasing nitrogen retention and reducing ureagenesis and amino acid oxidation. To determine whether the improvement in amino acid conservation with ST treatment involves regulation of urea cycle enzyme activities in both liver and intestine, growing swine were treated with either ST (150 microg x kg(-1) x d(-1)) or saline for 7 d. Fully fed pigs (n = 20) were infused intravenously for 2 h with NaH(13)CO(3) followed by a 4-h intraduodenal infusion of [1-(13)C]phenylalanine. Arterial and portal venous blood and breath samples were obtained at baseline and steady-state conditions for measurement of amino acid and blood urea nitrogen (BUN) concentrations and whole-body phenylalanine oxidation. Urea cycle enzyme activities were determined in liver and jejunum. ST decreased BUN (-46%), arterial (-34%) and portal venous (-43%) amino acid concentrations and whole-body phenylalanine oxidation (-30%). The activities of carbamoylphosphate synthase-I (-45%), argininosuccinate synthase (-38%), argininosuccinate lyase (-23%), arginase (-27%), and glutaminase (-18%), but not of ornithine carbamoyltransferase, ornithine aminotransferase, or glutamate dehydrogenase were reduced in liver of ST-treated pigs. ST slightly increased intestinal activity of glutaminase (+9%) but did not affect that of any other enzymes. ST decreased hepatic, but increased jejunal, N-acetylglutamate (an essential allosteric activator of carbamoylphosphate synthase-I; -26% and +32%, respectively) and carbamoylphosphate (a substrate for ornithine carbamoyltransferase; -20% and +28%, respectively) content. These results demonstrate that the reduced amino acid catabolism with ST treatment in growing pigs involves a reduction in hepatic urea cycle enzyme activities. The effect of ST treatment on porcine urea cycle enzymes is tissue-specific and is associated with a reduction in substrate availability for hepatic ureagenesis.

Ammonia detoxification and localization of urea cycle enzyme activity in embryos of the rainbow trout (Oncorhynchus mykiss) in relation to early tolerance to high environmental ammonia levels.

The present study investigated the role of ammonia as a trigger for hatching, mechanisms of ammonia detoxification and the localization of urea cycle enzymes in the early life stages of freshwater rainbow trout (Oncorhynchus mykiss). The key urea cycle enzyme carbamoyl phosphate synthetase III was found exclusively in the embryonic body (non-hepatic tissues); related enzymes were distributed between the liver and embryonic body. 'Eyed-up' trout embryos were exposed either acutely (2h) to 10 mmol l(-1) NH(4)Cl or chronically (4 days) to 0.2 mmol l(-1) NH(4)Cl. Time to hatching was not affected by either acute or chronic NH(4)Cl exposure. Urea levels, but not ammonia levels in the embryonic tissues, were significantly higher than in controls after both acute and chronic NH(4)Cl exposure, whereas there were no significant changes in urea cycle enzyme activities. Total amino acid levels in the embryonic tissues were unaltered by chronic ammonia exposure, but levels of most individual amino acids and total amino acid levels in the yolk were significantly lower (by 34-58%) than in non-exposed controls. The data indicate that trout embryos have an efficient system to prevent ammonia accumulation in embryonic tissue, by conversion of ammonia to urea in embryonic tissues and through elevation of ammonia levels in the yolk.