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PubMed1978

Lymph and intrarenal venous blood as indicators of canine renal cortical function.

Samplings of canine capsular and hilar lymph, and intrarenal venous blood were investigated as in vivo techniques for detecting renal cortical function in control and NH4Cl pretreated anesthetized animals. The glucose concentration of capsular, but not hilar, lymph was indicative of cortical drainage in both groups. When lymph flow was increased by elevating renal venous pressure, capsular lymph glucose concentration reflected cortical function only in the NH4Cl animals. The glucose concentration of intrarenal venous blood indicated cortical drainage in both the control and NH4Cl animals but the cortical function of a high para-aminohippuric acid extraction was only detectable in the NH4Cl group. Conclusions are that sampling of capsular lymph and intrarenal venous blood may prove to be useful techniques for studying certain in vivo cortical functions but the intrarenal venous technique is less sensitive than capsular lymph collection.

Administration, Oral↗

PubMed1988

Direct and simultaneous high-performance liquid chromatographic assay for the determination of p-aminobenzoic acid and its conjugates in human urine.

Procedures based on high-performance liquid chromatography (HPLC) were developed for identifying and measuring p-aminobenzoic acid (PABA) and its conjugate metabolites in human urine after oral doses of PABA. p-Aminohippuric acid (PAH), PABA, p-acetamidohippuric acid (PAHA) and p-acetamidobenzoic acid (PADB) in urine were resolved and determined by HPLC simultaneously and directly without extraction. A mobile phase consisting of 3% (v/v) acetonitrile in distilled water containing 0.005 M 1-heptanesulphonic acid in glacial acetic acid (PIC-B7) at pH 3.3 was eluted at 1 ml/min through a C18 Spherisorb column, followed by UV detection at 280 nm. After hydrolysis of urine samples at 37 degrees C for 3 h with beta-glucuronidase, the amounts of PABA-glucuronide and PADB-glucuronide were also determined. The retention times of PAH, a dominant peak which disappeared after hydrolysis, PABA, DABA (3,5-diaminobenzoic acid, as the internal standard), PAHA and PADB were 11.8, 14, 15, 18, 24 and 46 min, respectively. The 24-h urinary recoveries of PAH, PAHA, PADB, PADB-glucuronide, PABA and PABA-glucuronide after separate oral doses of 200 and 800 mg of PABA in one healthy subject were 43.4 and 48.1, 7 and 29.1, 11.2 and 11.8, 34.8 and 6.6, 0.2 and 0.3, and 1.0 and 2.4%, respectively. It is interesting that at high dose (800 mg) saturation of glucuronidation of PADB (N-acetylated PABA) appeared to occur, which resulted in an increase in the formation of PAHA, the glycine conjugate of PADB. Over 90% of the oral dose was accounted for by 8 h after administration.

4-Aminobenzoic Acid↗

PubMed1985

Clearance of compounds of different molecular size in the human placenta in vitro.

The placental clearance of selected water-soluble molecules and antipyrine was evaluated using a dual perfusion of human placental lobules in vitro. When graded according to decreasing clearance, the sequence of the molecules was as follows: water = antipyrine greater than urea greater than p-aminohippuric acid greater than inulin greater than dextran 20 greater than dextran 70. A close correlation was obtained between the clearance and the free diffusion coefficient of these molecules. However, no restricted diffusion was observed in the molecular range studied. These results suggest an equivalent pore size larger than 90 A for the human placental membrane in vitro.

Aminohippuric Acids↗

PubMed1984

PAH excretion in two species of cancroid crab Cancer irroratus and C. borealis.

To determine the characteristics of p-aminohippuric acid (PAH) excretion in Cancer irroratus and C. borealis, crabs were given a single injection of radiolabeled PAH and polyethylene glycol (PEG), and serum, urine, tissue, and seawater concentrations of each were measured over the next 6 days. In both species, PEG clearances averaged 5-10 ml serum/day; serum PAH concentrations fell exponentially after a 1-day equilibration period. In C. irroratus, urine PAH rose over the first 2 days and then fell; during days 2-6, mean urine PAH averaged two to four times mean serum PAH. In C. borealis, urine-to-serum ratios for PAH were 20-40 during days 2-6. PAH-to-PEG clearance ratios exceeded unity in both species, with ratios in C. borealis about three to six times those in C. irroratus. Tissue distribution data showed that C. irroratus midgut gland accumulated up to 50% of injected PAH, with gland-to-serum concentration ratios of 15-45; in this species, stomach fluid-to-serum PAH concentration ratios averaged 87, and nonurinary routes accounted for more than half of excreted PAH. In C. borealis, midgut gland-to-serum ratios never exceeded 2, and the urinary routes accounted for all excreted PAH. Thus, in C. irroratus, injected PAH accumulated in the midgut gland, with slow excretion through renal and nonrenal routes; in C. borealis, PAH remained in hemolymph, and only the renal route of excretion was utilized.

Aminohippuric Acids↗

PubMed1982

PAH transport in rock crab urinary bladder. II. Luminal and serosal steps.

To characterize the organic anion transport properties of the luminal (1) and serosal (s) membranes of crab urinary bladder, initial (10 min) fluxes (J) and tissue accumulations (Ac) of 10 microM PAH were measured in the absence and presence of 1 mM BCG (nontransported, high-affinity competitor). Control bladders exhibited net reabsorptive transport with a mean Jl leads to s/Js leads to 1 of about 7; concentrative transport of p-aminohippuric acid (PAH) occurred only across the luminal cell (c) membrane. With luminal PAH, luminal bromocresol green (BCG) reduced Jl leads to c, Jc leads to s, and Ac by about 85%; serosal BCG reduced Jc leads to s, increased Ac but had no effect on Jl leads to c. With serosal PAH, serosal BCG reduced Js leads to c, Jc leads to l, and Ac; luminal BCG had no effects on either fluxes or tissue accumulation. When bladder sheets were loaded with radiolabeled PAH, mounted in a chamber, and exposed to flowing crab Ringer solution on both sides, Jc leads to s was nearly twice as large as Jc leads to l; BCG significantly reduced Jc leads to s but not Jc leads to l. With unlabeled PAH in the efflux media, Jc leads to s was increased. The data are consistent with a model featuring an inwardly directed pump at the luminal membrane, a facilitated carrier at the serosal membrane, and nonmediated pathways at both membranes.

Aminohippuric Acids↗

PubMed1982

PAH secretion in the urinary bladder of a crab Cancer borealis.

Uptake of 10 microM p-aminohippuric acid (PAH) by sections of Cancer borealis urinary bladder was concentrative, saturable (Km 67 microM, Vmax 1.7 nmol.mg tissue-1.h-1), inhibitable by other organic anions, and dependent on medium Na and glycolytic metabolism. Bladders mounted in flux chambers exhibited net secretory transport of PAH, with serosa-to-lumen fluxes (Js leads to l) being about 4 times lumen-to-serosa fluxes (Jl leads to s). In 60-min flux chamber studies, tissue-to-medium ratios exceeded unity with serosal, but not luminal, PAH. Initial (10 min) fluxes and tissue accumulations (Ac) were measured in the absence and presence of 1-5 mM BCG (bromocresol green; competitor organic anion). With serosal PAH, serosal BCG (1 mM) reduced serosa-to cell flux (Js leads to c), Ac, and Jc leads to l by 60-75%. With luminal PAH, luminal BCG (1 mM) had no effect on Jl leads to c, Ac, or Jc leads to s; increasing the luminal BCG concentration to 5 mM reduced Jl leads to c, Ac, and Jc leads to s by 40-50%. The data are consistent with a model featuring an inwardly directed pump on the serosal membrane, cellular accumulation, and a facilitated carrier on the luminal membrane.

Aminohippuric Acids↗

PubMed1988

Coupled transport of p-aminohippurate by rat kidney basolateral membrane vesicles.

p-Aminohippuric acid (PAH) transport by basolateral membrane (BLM) vesicles isolated from rat renal cortex was stimulated very little by a Na+ gradient (out greater than in). However, when micromolar concentrations of glutaric acid or alpha-ketoglutaric acid were added in the presence of a out greater than in Na+ gradient, PAH uptake was accelerated greater than 20-fold and an overshoot of greater than fivefold was produced. Other anions, e.g., fumarate, stimulated PAH uptake very modestly under these conditions (approximately 2-fold), and that stimulation was totally prevented by short circuiting, i.e., with K+ (in = out) and valinomycin. Glutarate-stimulated uptake was inhibited by 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS) and probenecid and was slightly stimulated by the imposition of an inside-negative membrane potential. Furthermore, even in the absence of a Na+ gradient, glutarate-loaded vesicles exhibited a marked acceleration of PAH uptake (5-fold) and a modest overshoot (2.5-fold). These results suggest an indirect coupling of BLM PAH uptake to the Na+ gradient by a cyclic accumulation (Na+-dependent) of glutarate followed by its efflux from the vesicle in exchange for PAH. This coupled system was absent in apical membranes. Thus net secretory transport of PAH may entail Na+-dependent, glutarate-driven PAH uptake at the BLM, followed by the exit of PAH into the lumen down its electrochemical gradient, probably in exchange for other anions, e.g., Cl-, HCO3-, or OH-.

Aminohippuric Acids↗

PubMed1982

Choline and PAH transport across blood-CSF barriers: the effect of lithium.

The components of the blood-CSF barrier responsible for the transport of p-aminohippuric acid (PAH) and choline from CSF to blood were identified using in vitro preparations of frog choroid plexus and arachnoid membranes. Choline was transported out of CSF across the arachnoid, while PAH was transported out across the choroid plexus. Probenecid and ouabain blocked both processes. The effect of Li on these transport processes was tested by the addition of 5 mM LiCl to the incubation media. Li increased, by a factor of two, choline transport across the arachnoid, but there was no effect of Li on PAH transport across the plexus. Lithium was passively transported across the choroid plexus, and we suggest that the major transport pathway is through the tight junctions. The steady-state distribution of Li between the choroidal epithelium and the incubation medium was only half that expected for passive distribution. This suggests the existence of sodium/lithium countertransport in these epithelial cell membranes.

Aminohippuric Acids↗

PubMed1988

Organic anion transport by basal-lateral membranes: effect of PAH and furosemide on each other's transport.

The transport of organic anions by the kidney has been shown to be a carrier-mediated process. In an effort to learn more about this process, and examine the potential for two organic anions to compete for the same carrier site, studies were done which involved the transport of p-aminohippuric acid (PAH) and furosemide by vesicles made from basal-lateral membranes of rabbit kidney proximal tubules. Basal-lateral membranes were prepared by differential and ultracentrifugation. The transport was measured by using radiolabelled (3H) organic anions. The transport of each molecule was inhibited by probenecid, indicating that the carrier-mediated process for organic anion transport was functional in these studies. The results indicate that transport of PAH can be inhibited by furosemide in a concentration-dependent manner. This may indicate competition for the same carrier site. Inhibition of furosemide transport by PAH was not significant, perhaps due to much variability in the data. This variability may be due to nonspecific binding of furosemide to the vesicle, higher affinity of furosemide than of PAH for the receptor, or to the presence of more transport carriers for furosemide than for PAH. Experiments were done to determine the extent of nonspecific binding of furosemide. The results show that nonspecific binding of furosemide is extensive, indicating that this may contribute to the differences seen in the inhibition of transport. The data suggest that PAH and furosemide are transported by a common carrier-mediated process in the proximal tubule of the rabbit kidney.

Aminohippuric Acids↗

PubMed1985

Mechanism of renal concentration of technetium-99m glucoheptonate.

Seventy female Sprague-Dawley rats were studied to determine the mechanism of tubular localization and the effects of commonly encountered changes in hydration and acid-base balance on renal uptake and urinary excretion of technetium-99m glucoheptonate ([99mTc]GHA). The in-vivo protein binding and protein-free plasma clearance of [99mTc]GHA also were quantitated. Twenty additional rats were studied to determine the effects of PAH competition and probenecid blockade on renal uptake of [99mTc]dimercaptosuccinic acid (DMSA) in comparison with their effects on [99mTc]GHA localization. Kidney uptake of [99mTc]GHA averaged 11.17 +/- 0.49 (s.e.)% of the injected dose in control animals. This varied slightly among groups but was significantly reduced by probenecid blockade and para-aminohippuric acid (PAH) competition to 4.08 +/- 0.55 (p less than 0.005) and 2.39 +/- 0.14 (p less than 0.005), respectively. Technetium-99m DMSA was not affected in its renal accumulation by these maneuvers. The total plasma clearance of [99mTc]GHA was lower than iodine-125(125I)iothalamate but the clearance of the protein free supernate was higher, raising a possibility of some tubular secretion. Acidification of the urine which has been shown to reduce [99mTc]DMSA uptake appeared to have no effect on [99mTc]GHA. Hepatic uptake was minimal in all groups averaging less than 1% injected dose. These data demonstrate that renal accumulation of [99mTc]GHA is blocked by probenecid and PAH suggesting that it is actively concentrated in the proximal tubule by enzyme systems similar to those involved in PAH and hippuran transport. It appears that [99mTc]GHA uptake measures a different aspect of kidney function than [99mTc]DMSA.

Acid-Base Imbalance↗

PubMed2000

Regulation of mOAT-mediated organic anion transport by okadaic acid and protein kinase C in LLC-PK(1) cells.

Organic anion transporters in the kidney proximal tubule play an essential role in eliminating a wide range of organic anions including endogenous compounds, xenobiotics, and their metabolites, thereby preventing their potentially toxic effects within the body. We have previously cloned a cDNA encoding an organic anion transporter from mouse kidney (mOAT) (Lopez-Nieto, C. E., You, G., Bush, K. T., Barros, E. J. G., Beier, D. R., and Nigam, S. K. (1997) J. Biol. Chem. 272, 6471-6478; Kuze, K., Graves, P., Leahy, A., Wilson, P., Stuhlmann, H., and You, G. (1999) J. Biol. Chem. 274, 1519-1524). In the present study, we assessed the potential for regulation of this transporter by heterologous expression of mOAT in the pig proximal tubule-like cell line, LLC-PK(1). We report here that both protein phosphatase (PP1/PP2A) inhibitor, okadaic acid, and protein kinase C (PKC) activators down-regulate mOAT-mediated transport of para-aminohippuric acid (PAH), a prototypic organic anion, in a time- and concentrationdependent manner. However their mechanisms of action for this down-regulation are distinct. Okadaic acid modulated PAH transport, at least in part, through phosphorylation/dephosphorylation of mOAT; phosphoamino acid analysis indicated this phosphorylation occurs on serine. In contrast, PKC activation induced a decrease in the maximum transport velocity (V(max)) of PAH transport without direct phosphorylation of the transporter protein. Together these results provide the first demonstration that regulation of organic anion transport by mOAT is likely to be tightly controlled directly and indirectly by phosphatase PP1/PP2A and PKC. Our results also suggest that kinases other than PKC are involved in this process.

Animals↗

PubMed1987

p-Aminohippurate transport in basal-lateral membrane vesicles from rabbit renal cortex: stimulation by pH and sodium gradients.

p-Aminohippuric acid (PAH) uptake was studied in basal-lateral membrane vesicles prepared from rabbit renal cortex. An outwardly directed hydroxyl gradient (pHo = 6.0, pHi = 7.6) stimulated PAH uptake slightly over that when the internal and external pH values were equal at 7.6. A 100 mM sodium gluconate gradient directed into the basal-lateral membrane vesicles increased PAH uptake about 2-fold over that when N-methyl-D-glucamine or potassium gluconate gradients were present. When hydroxyl and sodium gradients were simultaneously imposed (pHo = 6.0, pHi = 7.6 and 100 mM sodium gluconate extravesicularly) PAH uptake was stimulated greater than with the pH or Na+ gradient alone. In fact, an 'overshoot' was observed. Countertransport experiments showed that either intravesicular PAH or intravesicular PAH and Na+ could stimulate 3H-PAH uptake. Probenecid, an inhibitor of organic anion transport, inhibited both the hydroxyl-stimulated and Na+ gradient-stimulated PAH uptake but the greatest inhibition by probenecid was seen when the hydroxyl and sodium gradients were both present. Thus, it is proposed that the driving force for PAH accumulation across the basal-lateral membrane of the proximal tubule is a transport system which moves Na+ and PAH into the cell for an hydroxyl ion leaving the cell, i.e. a sodium-dependent anion-anion exchange system.

Aminohippuric Acids↗

PubMed1985

Maturation of Rb+ and PAH accumulation by rabbit anterior uvea and choroid plexus.

In vitro accumulation of radioactive para-aminohippuric acid (3H-PAH) and rubidium (86Rb+) by the anterior uvea, ciliary processes, and the choroid plexus was evaluated in tissues from newborn and various aged rabbits. Accumulation of PAH was present in the anterior uvea at 1 day of age (tissue to media ratio, T/M, of 2.1 +/- 0.2) and remained at this level for the first 14 days of life. Accumulation did not rise to adult levels until 21 days of age (T/M 5.5 +/- 0.6). Rubidium accumulation in the anterior uvea, a measure of Na+, K+-pump activity, was higher than adult values 6 hr after birth (T/M25.2 +/- 0.9). Activity remained elevated through day 28 and did not fall to adult levels until day 60 (T/M 13.4 +/- 0.6). Accumulation studies on isolated ciliary processes were similar to those obtained from anterior uveal tissue. Daily subcutaneous injections of penicillin (300,000 units/kg/day) for 1 week had no effect on anterior uvea PAH accumulation (penicillin T/M was 1.7 +/- 0.1 and saline control T/M was 2.0 +/- 0.2). Accumulation of either 3H-PAH or 86Rb+ by the choroid plexus was present 1 day after birth in amounts that were similar to adult values and did not change during the 90 days of testing.

Aminohippuric Acids↗

PubMed1998

Pitfalls in measuring inulin and para-amino-hippuric acid clearances.

Various carbohydrates and a variety of widely used medicines interfere with the generally used laboratory methods for determining inulin and para-aminohippuric acid (PAH). When these pitfalls are not recognized, false measurements of inulin and PAH clearances, which represent glomerular filtration rate and renal plasma flow respectively, are obtained. When performing these tests a careful history of dietary habits and oral drug therapy must be taken.

Ammonium Chloride↗

PubMedDate not supplied

Whole blood and plasma amino acid transfers across the portal drained viscera and liver of the pig.

Whole blood (WB) and plasma (P) amino acid transfers across the portal drained viscera and the liver were determined during 6 h of a constant p-aminohippuric acid infusion in three hourly-fed Landrace x Large-White pigs (30.5 kg, mean live weight) surgically prepared with chronically inserted catheters in a mesenteric vein (MV), the portal vein (PV), an hepatic vein (HV) and the carotid artery (CA). Plasma and WB amino acid concentrations were determined in the CA, PV and HV. The plasma/WB ratios showed no significant differences for vessels except for lysine and glutamate for which this ratio is significantly higher in the HV and in the PV for lysine. This suggests that the PV lysine and HV glutamate were preferentially transported in the plasma. In the PV, threonine, valine and alanine are transported by both plasma and red blood cells. These data show that the contribution of plasma and whole blood to amino acid transport can be different between amino acids and between individual tissues.

Alanine↗

PubMed1975

On the problem of a common hepatic transport process for steroids. Uptake of 3-H-taurocholic acid and 3-H-corticosterone into rat liver slices.

The uptake of 3-H-oubain (2 times 10- minus 9 and 2 times 10- minus 5 M), 3-H-taurocholic acid (2 times 10- minus 8 and 2 times 10- minus M) and 3-H-corticosterone (2 times 10- minus 9 and 2 times 10- minus 5M) into rat liver slices was investiged. The steroids were extensively accumulated as demonstrated by the following T/M ratios: ouabain: 3.82 and 3.72 resp.; taurocholic acid: 10.49 and 9.88 resp.; corticosterone: 12.84 and 8.41 resp.. Only corticosterone uptake was dependent on the medium concentration but without alterations under anaerobic conditions. The contrary could be found in case of ouabain and taurocholic acid. Further differences between the steroids investigated could be shown in experiments on the interaction with organic anions (probenecid, para-aminohippuric acid, citrate and alpha-ketoglutarate). At least no competition among the steroids concerning their uptake into liver tissue could be demonstrated. The results are discussed in contradiction to the current theories concerning the hepatic uptake of steroid compounds.

Aminohippuric Acids↗

PubMed1982

[Elimination kinetics following repeated administrations of chemotherapeutic agents].

Repeated administrations of the chemotherapeutic agents nalidixic acid, retacillin, oxacillin, ampicillin and gentamicin acts on the transport of organic acids by the kidney. With the exception of nalidixic acid, all the chemotherapeutic agents under study produced a significant dose-dependent increase in the renal clearance of p-aminohippuric acid (PAH) in adult rats aged 55 days. Due to nephrotoxic effects, the renal clearance of PAH is decreased by very high doses of nalidixic acid and gentamicin. In contrast to this, the changes in PAH clearance were less marked in 10-day-old rats in which the ability of the kidney to transport organic acids is low. On repeated administrations of chemotherapeutic agents, one has on principle to consider the production of a diminishing effect due to increased renal clearance.

Animals↗

PubMed1998

Renal arginine metabolism in fasted rats with subacute short bowel syndrome.

1. Arginine can be produced in the kidney from citrulline. An important source of circulating citrulline is the intestinal breakdown of glutamine. Consequently, partial enterectomy leads to decreased plasma citrulline levels. The aim of the present study was to investigate the effect of diminished arterial citrulline levels on renal arginine production and total-body free arginine pools.2. Renal amino acid metabolism was studied 24 h after 75% small bowel resection in rats fasted overnight (16 h) (n=12; total fast 40 h). Sham-operated (n=9) and non-operated 16-h and 40-h fasted controls were studied in parallel (n=8/n=7). During anaesthesia, L-(2, 3-3H)-arginine and para-aminohippuric acid were infused until steady state. Subsequently, arterial and renal venous blood samples were taken. Concentrations of para-aminohippurate and amino acids and specific activity of arginine and citrulline were measured to calculate renal plasma flow, net renal uptake or release, and unidirectional influx or efflux of arginine and citrulline, as well as whole-body arginine turnover.3. Arterial citrulline was decreased in enterectomized rats compared with sham-operated rats (23+/-3 versus 44+/-6 microM). Net renal citrulline uptake and arginine release were almost stoichiometric (-36+/-7 and 38+/-6 nmol.min-1. 100 g-1 body weight respectively in sham-operated rats) and were both diminished by 50% in enterectomized versus sham-operated rats. In all groups, net renal arginine production accounted for less than 10% of whole-body rate of arginine appearance (488 nmol.min-1.100 g-1 body weight in the sham group). Despite decreased net renal citrulline consumption and renal arginine production in enterectomized rats, whole-body rate of arginine appearance and arterial arginine did not change significantly.4. In conclusion, net renal arginine production is reduced 24 h after 75% enterectomy in fasted rats. However, this does not have important effects on whole-body arginine production.

Amino Acids↗