Urinary fluorescent metabolite O-aminohippuric acid is a useful biomarker for lung cancer detection.

INTRODUCTION: Urine contains diagnostically important metabolites that can act as natural fluorophores. However, whether these fluorescent metabolites can be used in lung cancer diagnosis is unknown. OBJECTIVES: This study was conducted to determine whether fluorescent urinary metabolites could be useful biomarkers for lung cancer detection. METHODS: A total of 46 lung cancer patients and 185 volunteers without cancer were evaluated between November 2013 and November 2014. Samples of the first urine of the day were collected from lung cancer patients and diagnosed at the Hamamatsu University School of Medicine and the Hamamatsu Medical Center prior to cancer treatment, and from volunteers without cancer at the Hamamatsu Medical Imaging Center. Fluorescent urinary metabolites were screened by high-performance liquid chromatography and select effective fluorescent substances for distinguishing cancer from non-cancer status. RESULTS: The fraction of patients at each stage of cancer severity were: 41.3% stage I, 8.7% stage II, 19.6% stage III, and 30.4% stage IV. A robust predictive biomarker for lung cancer was selected by the multivariate logistic analysis of fluorescent metabolites and identified to be O-aminohippuric acid (OAH). The area under the curve (AUC) data for OAH was 0.837 (95% CI 0.769-0.898, P < 0.001). CONCLUSION: We identified a fluorescent urinary metabolite that can predict lung cancer. OAH exceeds the AUC (0.817) of lung cancer detection by AminoIndex® cancer screening, can be analyzed non-invasively without additional sample processing, and may be a valuable addition to existing lung cancer prediction models.

Human organic anion transporter OAT1 is not responsible for glutathione transport but mediates transport of glutamate derivatives.

Due to their clearance function, the kidneys are exposed to high concentrations of oxidants and potentially toxic substances. To maintain cellular integrity, renal cells have to be protected by sufficient concentrations of the antioxidant glutathione (GSH). We tested whether GSH or its precursors are taken up by human organic anion transporters 1 (OAT1) and 3 (OAT3) stably expressed in HEK293 cells. GSH did not inhibit uptake of p-aminohippurate (PAH) or of estrone sulfate (ES) in OAT3-transfected HEK293 cells. In OAT1-transfected cells, GSH reduced the uptake of PAH marginally. Among the GSH constituent amino acids, glutamate, cysteine, and glycine, only glutamate inhibited OAT1, but labeled glutamate was not taken up by a probenecid-inhibitable transport system. Thus OAT1 binds glutamate but is unable to translocate it. The GSH precursor dipeptide, cysteinyl glycine (cysgly), and the glutamate derivative N-acetyl glutamate (NAG), inhibited uptake of PAH when present in the medium and trans-stimulated uptake of PAH from the intracellular side, indicating that they are hitherto unrecognized transported substrates of OAT1. N-acetyl aspartate weakly interacted with OAT1, but aspartate did not. NAG inhibited also OAT3, albeit with much lower affinity compared with OAT1, and glutamate did not interact with OAT3 at all. Taken together, human OAT3 and OAT1 cannot be involved in renal GSH extraction from the blood. However, OAT1 could support intracellular GSH synthesis by taking up cysteinyl glycine.

The functional capacity of hypertrophied nephrons. Effect of partial nephrectomy on the clearance of inulin and PAH in the rat.

A method is described for the measurement of inulin and PAH clearances in rats without killing the animal. Inulin clearance measurements of partially nephrectomized rats and sham-control rats were made before operation and at intervals following the operation; inulin clearances were determined on normal rats at intervals during their growth. In another series of partially nephrectomized rats and their sham-operated controls, inulin and PAH clearances were determined in all the animals before and at intervals following the operation. Glomerular counts were made in some rats. After partial nephrectomy the inulin clearance is reduced but not as much as would be expected considering the amount of renal tissue removed. The mean inulin clearance per nephron is greatly increased in the partially nephrectomized rat when compared to the value determined for the control rat. The inulin clearance of the partially nephrectomized rat shows a progressive decline which is first clearly evident about 6 months after operation. The sham-operated rats showed an inulin clearance slightly less than that of nonoperated controls about 24 to 30 wk after the operation. In the normal rats the inulin clearance relative to body weight is much greater in rats with a mean weight of 197 +/- 3 g than in normals which are older and heavier. The PAH dearance of the partially nephrectomized rat is reduced following operation but undergoes no further change in the ensuing 24 wk.

Evaluation of a stroma-free hemoglobin solution for use as a plasma expander.

The preparation of large quantities of a stable, stroma-free hemoglobin solution without coagulant activity is described. Following infusion of this solution into phlebotomized dogs, there is no methemoglobin formation, no adverse effects on vital signs, and no demonstrable activation of blood coagulation. The hemoglobin maintains its oxygen-carrying capacity and liberates oxygen into tissues. Acute and chronic effects on renal function following infusion of this preparation were also studied and no effect on clearance of urea, creatinine, or P.A.H. could be demonstrated. There was no change in urinary output and histological sections revealed no lesions attributable to hemoglobin toxicity. It is concluded that a stroma-free hemoglobin solution may have use as a plasma expander.

The intrarenal distribution of tritiated para-aminohippuric acid determined by a modified technique of section freeze-dry radioautography.

Section freeze-dry radioautography has been used to examine the intrarenal distribution of a water-soluble organic acid (para-aminohippuric acid (PAH-3H)) under constant-infusion, steady-state conditions in mouse and rat kidney in vivo. The technique described here has the following advantages: (a) Sectioning and freeze-drying are accomplished in a closed cryostat at temperatures below -40 degrees C; (b) Handling of the section is facilitated by mounting of the section-to-be on adhesive-coated Saran Wrap prior to cutting; (c) Unembedded freeze-dried sections are attached to photographic film at ambient temperature in the dark room; (d) Fixation follows completion of radioautographic exposure and precedes photographic development; (e) Permanent close contact is maintained between tissue and film. Morphologic preservation compared favorably with that obtained by optimal fixation techniques, which, however, permit diffusion. Cellular accumulation of PAH-3H during secretion was demonstrated in the proximal tubule under steady-state conditions in vivo. The cellular concentration of PAH-3H was uniform throughout the length of the proximal tubule in mouse and rat kidney.

Maturation of renal organic acid transport: substrate stimulation by penicillin.

Renal p-aminohippurate transport in rabbits increased rapidly from birth to 4 weeks of age and then declined to that observed in adults. Penicillin administration to pregnant does or newborn rabbits stimulated the developing transport system, but did not increase the peak observed at 4 weeks. Therefore the continued presence of substrate (penicillin) during development significantly enhances the rate of maturation.

Urate-2-14C transport in the rat nephron.

Intrarenal transport of urate-2-(14)C was studied in anesthetized rats using the microinjection technic. During saline diuresis, small volumes of urate-2-(14)C (0.24-0.48 mM) and inulin-(3)H were injected into surface proximal and distal convoluted tubules, and ureteral urine was collected serially. Total (74-96%) and direct (57-84%) urate recovery increased significantly the more distal the puncture site. Delayed recovery (+/-20%) remained approximately the same regardless of localization of the microinjection. After proximal injections, total and direct recoveries of urate-2-(14)C were significantly higher in rats treated with probenecid, pyrazinoate, or PAH than during saline diuresis alone, while the excretion rates were comparable after distal injection. Delayed recovery was not altered by drug administration. The decreased proximal reabsorption of urate is presumably due to an effect of the drugs on the luminal membrane of the nephron. For perfusion at high urate concentrations, nonradioactive urate was added to the injectate (0.89-1.78 mM). Urate-2-(14)C recovery was almost complete and there was no delayed excretion, demonstrating saturation kinetics. These findings are compatible with a carrier-mediated mechanism for urate transport probably located at the luminal border of the proximal tubular epithelium. No definitive evidence for urate secretion was found in these studies.

A natriuretic factor in the serum of patients with chronic uremia.

Sera from chronically uremic and normal individuals were subjected to gel filtration with Sephadex G-25 and the same fraction of both was infused into rats with a decreased nephron population to determine the effects on sodium excretion. Sodium excretion rate and fractional sodium excretion increased slightly with the normal fractions; but the increase in both functional parameters produced by the uremic fractions was substantially and significantly greater. The natriuresis could not be explained by associated changes in glomerular filtration rate (GFR), para-aminohippurate (PAH) clearance, filtration fraction, hematocrit, or blood pressure. The possibility thus exists that the inhibitor affected some component part of the transepithelial sodium transport system. The elution characteristics of the fraction plus certain of its physicochemical properties suggest that the inhibitor of sodium reabsorption by the rat nephron may be identical with the inhibitor of PAH uptake by kidney slices and the inhibitor of transepithelial sodium transport by the frog skin and toad bladder previously found in the serum of chronically uremic patients.

Role of the sympathetic nervous system in the renal response to hemorrhage.

In 12 studies, a femoral artery and vein of a donor dog treated with desoxycorticosterone were connected by tubing to a renal artery and vein of a recipient dog treated with desoxycorticosterone, and the kidney with its nerve supply intact was perfused at femoral arterial pressure. Infusion of normal saline, which contained albumin, from 2.7 to 3.1 g/100 ml, in the donor produced significant natriuresis in a kidney of the donor (from 112 to 532 muEq/min) and in the perfused kidney (from 60 to 301 muEq/min) of the recipient. Increased sodium excretion in the perfused kidney was associated with an increase in the clearances of inulin and para-aminohippurate (P < 0.01) and a decrease in hematocrit of perfusing blood (P < 0.01). Infusion was continued in the donor while recipient was bled 23 ml/kg, with a decrease in mean arterial pressure from 152 to 130 mm Hg. Sodium excretion in perfused kidney decreased from 301 to 142 muEq/min (P < 0.01), whereas sodium excretion in donor was unchanged (506 VS. 532 muEq/min; P > 0.3). Clearance of inulin by perfused kidney was not significantly affected by bleeding (26 +/-SE 2 VS. 25 +/-SE 2; P > 0.2), but the clearance of para-aminohippurate was decreased by bleeding (P < 0.01), so that filtration fraction increased. As the perfused kidney of the recipient dog continued to receive blood from the natriuretic donor dog when the recipient dog was bled, the decrease in sodium excretion that bleeding produced in the perfused kidney was presumably mediated by renal nerves. Thus, an increase in nervous stimuli to the kidney that is not sufficient to decrease glomerular filtration rate can increase the tubular reabsorption of sodium and thereby significantly decrease its excretion. This property of the sympathetic nervous system to affect tubular reabsorption of sodium suggests that an increase in sympathetic activity may constitute an important mechanism for the renal conservation of sodium when intravascular volume is contracted by hemorrhage or other cause.

Effects of potassium deficiency on renal function in the dog.

Serial determinations of the renal clearance for inulin and para-aminohippuric acid (PAH), maximum renal tubular reabsorptive rate for glucose, maximum urinary concentrating ability, total exchangeable potassium, extracellular volume, and plasma sodium and potassium concentrations were done in seven dogs before and after dietary potassium depletion. The same measurements were also made in two of the dogs during potassium repletion. Inulin and PAH clearances and transport maxima for glucose decreased progressively during depletion. These changes correlated well with both the duration of depletion and the extent of depletion as measured by total exchangeable potassium. Decreases in inulin and PAH clearance closely paralleled each other, suggesting that there might be a renal hemodynamic basis for both effects. The decreases in transport maxima for glucose were greater than those for inulin or PAH clearance, indicating the presence of a defect in the cellular transport mechanism for glucose. In the dogs that were repleted, renal function gradually returned to the predepletion state. No significant changes were found in extracellular volume or plasma sodium concentration during depletion. Renal concentrating ability decreased only moderately during depletion, with the decrease correlating better with plasma potassium concentration than with total exchangeable potassium. This finding contrasts with the marked decrease in concentrating ability and the severe polydipsia and polyuria found in animals depleted of potassium with the aid of corticosteroids. The results of the present study emphasize the importance of considering species differences and the method of producing depletion in interpreting studies of the effects of hypokalemia on renal function.

A micropuncture study of potassium excretion by the remnant kidney.

In order to study the mechanism of enhanced potassium excretion by the remaining nephrons of the remnant kidney, micropuncture and clearance experiments were carried out in rats after surgical ablation of 3/4 of the total renal mass. The potassium intake in all animals was approximately 5 meq/day. Animals were studied 24 h and 10-14 days after 3/4 nephrectomy. Balance measurements in the chronic animals before micropuncture study indicated that 24 h K(+) excretion by the remnant kidney was equal to that of the two kidneys before ablation of renal mass. Measurements of distal tubular inulin and potassium concentrations revealed progressive reabsorption of potassium in this segment of the nephron in both the 24-h and chronic 3/4-nephrectomized rats, as well as in normal control rats. A large increase in tubular fluid potassium content occurred between the end of the distal tubule and the final urine in the 3/4-nephrectomized rats, but not in the normal controls. These observations suggest that the segment of the nephron responsible for enhanced potassium excretion by remaining nephrons was the collecting duct. In additional experiments, potassium was completely eliminated from the diet of chronic 3/4-nephrectomized rats before micropuncture study. In these animals, no addition of K(+) occurred beyond the distal tubules. Normal rats infused with 0.15 M KCl to acutely elevate serum K(+) concentration, demonstrated reabsorption of K(+) in the distal tubule and a large addition of K(+) to the urine beyond the distal tubule. We conclude that the collecting duct is the major site of regulation of urinary potassium excretion in normal rats and is responsible for the adaptation to nephron loss by the remnant kidney.

Tubular reabsorption of sodium during acute and chronic volume expansion in man.

Renal hemodynamics and tubular fractional sodium reabsorption (FSR) were evaluated by clearance techniques during acute and chronic extracellular volume expansion in man. (1 - V/GFR) x 100 was used as an index of proximal and (C(H2O)/V) x 100 as an estimate of distal fractional reabsorption. After acute loading with isotonic saline 37 ml/kg body wt, proximal FSR decreased by 4.8% and distal FSR decreased by 4.4%. After comparable chronic expansion by mineralocorticoids ("escape"), proximal FSR also decreased by 3.9%, but distal reabsorption was not altered.In separate studies, subjects were progressively infused with saline to 57 (E(1)) and to 80 (E(2)) ml/kg body wt, and appeared to divide into "excreters" (maximum U(Na)V > 1000 muEq/min) and "nonexcreters" (maximum U(Na)V < 550 muEq/min). In the excreters, GFR rose, proximal FSR decreased by 7.1% after E(1) and only 0.9% further after E(2). Distal FSR fell by 14.8% after E(1) and by an additional 4.9% after E(2). In the nonexcreters, GFR was stable and proximal FSR did not fall significantly after E(1) or E(2). Distal FSR decreased 4.5% after E(1) and 1.3% further after E(2). It is concluded that both acute and chronic extracellular expansion decrease proximal FSR in man, but only acute loading depresses distal FSR. Ability of some men to excrete sodium rapidly after acute infusion is related to larger increases in GFR and greater decreases in both proximal and distal FSR than occur in men in whom natriuresis is more limited.

Functional adaptation of nephrons in dogs with acute progressing to chronic experimental glomerulonephritis.

Although a diminished fractional excretion of sodium (FENa) is the hallmark of acute proliferative glomerulonephritis (APGN), an enhanced natriuresis per glomerular filtration rate (GFR) in the chronic phases of this disease has been reported. We studied this adaptive response utilizing two different split-bladder dog models with unilateral, and a third group of dogs with bilateral Masugi's nephritis. Group I. Six dogs with unilateral nonaccelerated APGN studied a mean of 6 days after induction had a mean base-line APGN/intact kidney GFR of 31/50 ml/min (P less than 0.005) and FENa of 0.2/0.75% (P less than 0.005). Acute volume expansion caused a smaller absolute increase in FENa from the APGN kidney, 1.6%, than from the intact kidney, 4.0%, (P less than 0.01). Maximum tubular secretion of rho-aminohippuric acid/GFR (TmPAH/GFR) measured in three dogs was higher in the APGN kidney than intact kidney, 13.1 vs. 9.3 mg/dl. Subsequent studies on three of the six dogs when the disease had become chronic demonstrated a reversal in the pattern of sodium excretion in response to volume expansion. Group II. Six dogs with accelerated unilateral APGN (dogs presensitized to antibody source) studied a mean of 5 days after induction had a mean base-line APGN/intact kidney GFR of 16/57 ml/min and FENa of 0.22/0.12% (P less than 0.1). Contrary to group I, volume expansion caused a greater absolute increase in FENa from the APGN kidney, 5.8%, than from the intact kidney, 2.9% (P less than 0.05). TmPAH/GFR studied in four dogs was similar for both kidneys, 17.9 and 18.5 mg/dl for the APGN kidney and intact kidney, respectively. Group III. Sequential studies were performed on seven dogs with bilateral nonaccelerated APGN. Initially each demonstrated sodium retention and a smaller absolute increase in FENa in response to volume expansion compared to a predisease control study. With disease progression, volume expansion induced a greater absolute increase in FENa than in the control study. We concluded that (a) the fractional excretion of sodium from the APGN kidney will be less or greater than the contralateral intact kidney or control study depending on the severity and/or chronicity of the disease, possibly as the result of morphologic alterations; (b) the degree of extracellular fluid volume expansion is an important variable influencing similarity of glomerulotubular balance between the APGN and contralateral intact kidney; and (c) the "intact nephron hypothesis" applies in a limited fashion to kidneys with APGN in the absence of volume expansion just as it does for kidneys with chronic glomerulonephritis or pyelonephritis.

Effects of acute unilateral renal denervation in the rat.

Studies were undertaken to characterize the renal responses to acute unilateral renal denervation and the mechanisms involved in these responses. Denervation was produced in anesthetized nondiuretic rats by application of phenol to the left renal artery. Studies were also performed in sham-denervated nondiuretic rats. Whole kidney and individual nephron studies were performed before and after denervation or sham denervation. Denervation increased urine volume from the left kidney to about twice its control value (P less than 0.001) and increased urinary sodium excretion from 332 neq min minus -1 to 1,887 neq min minus -1 (P less than 0.001). Glomerular filtration rate (GFR) and renal plasma flow (RPF) remained unchanged in both kidneys after the procedure. The innervated right kidney showed no changes in urine volume or in sodium excretion. After denervation, late proximal ratio of tubular fluid inulin concentration to that of plasma [(F/P)In] decreased from 2.23 to 1.50 (P less than 0.001) while single nephron GFR remained unchanged. Absolute reabsorption decreased from 16.5 to 9.9 n. min minus -1 (P less than 0.001). (F/P)In ratios were also decreased in early distal (from 6.21 to 3.18, P less 0.001) and late distal convolutions (from 16.41 to 8.33, P less than 0.001) during the experimental period. (F/P)Na ratios remained unchanged in the early distal convolutions, but increased from 0.18 to 0.38 (P less than 0.01) in late distal convolutions after denervation. Absolute Na reabsorption after denervation increased in the loop of Henle, distal convolution, and collecting ducts. Any changes in intrarenal hydrostatic pressures after denervation were always small. There were no changes in GFR, RPF, urine volume, urinary sodium excretion, or late proximal (F/P)In after sham denervation. We conclude that the diuresis and natriuresis seen after acute renal denervation were caused by a marked depression of sodium and water reabsorption in the proximal tubule with partial compensation in more distal nephron segments. These responses appeared to be unrelated to systemic or intrarenal hemodynamic changes. The results demonstrate an effect of the renal nerves on proximal tubular function.

Uricosuric agents in uremic sera. Identification of indoxyl sulfata and hippuric acid.

Serum and urine from chronically uremic patients and normal individuals were subjected to gel filtration of Sephadex-G10. The effects of the eluted fractions on the uptake of urate and para-aminohippurate by isolated cortical tubules of rabbit kidney were investigated. According to the origin of the samples, one to three major groups of fractions inhibiting both urate and para-aminohippurate transport were disclosed. The first eluted group occurred for all the samples under study. The second one was demonstrated in both sera and urines from uremic patients but only in urines from normal individuals. The third one was exclusively detected in uremic sera and urines. Among all the compounds identified, only hippuric acid, eluted in the fractions of the second group, was capable of inhibiting the uptake of urate and para-aminohippurate in vitro. The concentration for which this inhbiitory effect of hippuric acid occurred was in the range of that existing in uremic sera. Indoxyl sulfate, which accumulates to very high concentrations in uremic serum, could not be disclosed in the above-mentioned fractions. This is explained by the strong adsorption of this indole derivative to Sephadex gel. Potassium indoxyl sulfate, when tested in vitro at the concentration existing in uremic serum, substantially inhibited the uptake of both urate and para-aminohippurate. In normal subjects, ingestion of hippuric acid or potassium indoxyl sulfate significantly increased fractional urinary excretion of uric acid. On the basis of these results, it is suggested that progressive retention of hippuric acid, indoxyl sulfate, and other yet unidentified inhibitors may explain the gradual increase in urinary fractional excretion of urate observed in uremia. The present results may be viewed as an example of a mechanism in which retention of normally excreted end products is responsible for adaptation of tubular transport in uremic subjects.

The relative contributions of reabsorptive rate and redistributed nephron filtration rate to changes in proximal tubular fractional reabsorption during acute saline infusion and aortic constriction in the rat.

The absolute rate of reabsorption by superficial rat proximal tubules was measured by the in situ microperfusion technique under conditions of hydropenia, infusion of saline, and infusion of saline plus aortic constriction sufficient to decrease whole kidney filtration rate below hydropenic levels. Fractional reabsorption was measured in adjacent filtering nephrons by collecting and recollecting tubular fluid from late proximal convolutions during each experimental condition. During hydropenia, the absolute rate of proximal tubular reabsorption averaged 3.56 +/-0.60 nl/min per mm and late proximal tubular fractional reabsorption averaged 0.56 +/-0.10. From these two measurements and measurements of tubule length to the site of micropuncture, a value for filtration rate was calculated for filtering nephrons. During hydropenia this value averaged 32.9 +/-7.1 nl/min. Saline infusion increased sodium excretion to 5.5% of the filtered load as the absolute rate of proximal tubular reabsorption decreased 38% and fractional reabsorption decreased 45%. Calculated superficial nephron filtration rate increased 21% which on the average was identical with the simultaneously measured increase in whole kidney filtration rate. Similar results were obtained in a separate group of animals by the technique of total collection of late proximal tubular fluid. Aortic constriction during saline infusion decreased whole kidney and calculated nephron filtration rate to the same degree and to values lower than those during hydropenia. Fractional reabsorption increased but not to hydropenic values. The persistent natriuresis during aortic constriction was associated with a continued depression of the absolute rate of proximal tubular reabsorption which was sufficient to maintain an increased delivery of filtrate out of the proximal tubule despite the fall in nephron filtration rate. These results indicate that depressed fractional reabsorption in the proximal tubule during acute saline infusion is due predominantly to a decrease in absolute reabsorptive rate and to a lesser extent to an increase in superficial nephron filtration rate which is proportional to the increase in whole kidney filtration. Continued natriuresis when filtration rate is decreased during saline infusion can be accounted for entirely by the persistent large reduction in the absolute rate of proximal tubular reabsorption.

Effects of acute volume expansion and altered hemodynamics on renal tubular function in chronic caval dogs.

It is well established that dogs with chronic partial constriction of the thoracic inferior vena cava develop sodium retention, ascites, and respond poorly to acute saline loading. A group of such chronic caval dogs, and a group of normal controls were studied during hydropenia, and again after acute saline loading by clearance and recollection micropuncture techniques. After volume expansion, the caval dogs excreted 52 muEq/min per kidney of sodium compared with 370 muEq/min per kidney for the normal controls. During hydropenia and after the saline infusions, single nephron filtration rates, fractional reabsorption of sodium within the proximal tubule, and proximal delivery of filtrate to the distal nephron were comparable in both groups of dogs. Micropuncture of distal tubular segments confirmed that the loop of Henle was the major site for salt and water retention in the expanded caval dogs. Alteration of intrarenal hemodynamics by vasodilating one kidney and elevating systemic arterial blood pressure induced a normal natriuretic response in the saline-loaded caval dogs. Proximal tubular function remained unchanged and the loop of Henle appeared to be the major site responsive to these hemodynamic maneuvers. These same experiments in saline-loaded control dogs had no effect on function of the proximal or distal nephron and did not increase urinary excretion of sodium or water. These experiments provide evidence that the loop of Henle is the major site for sodium retention in volume-expanded chronic caval dogs excreting minimal amounts of sodium.

Renal tubular transport of organic acids. Studies with oxalate and para-aminohippurate in the rat.

The renal handling of oxalate was examined by free-flow micropuncture, intratubular microinjection, and droplet precession techniques in the rat. After the sustained i.v. infusion of [(14)C]oxalate, the fractional delivery of oxalate from the early portions of the proximal tubule was 120.1+/-4.4%, indicating net secretion. Fractional delivery rates from the late proximal tubule (124.6+/-6.1), distal tubule (120.9+/-2.9), and final urine (126.2+/-2.9%) were not different from that of the early proximal tubule. Direct intratubular microinjections of oxalate into the early proximal tubule and late proximal tubule yielded urinary recovery rates of 85+/-3% and 101+/-2%, respectively, suggesting that oxalate absorption does occur in the mid-portions of the proximal tubule. Droplet precession studies confirmed a secretory flux for oxalate. In contrast to oxalate, para-aminohippurate (PAH), the more traditional marker for organic acid transport, was secreted in the late portions of the proximal tubule and in large measure at a site between the late proximal and distal tubules, presumably the pars recta. Probenecid inhibited PAH secretion but was without effect on net oxalate transport, oxalate absorption, or oxalate secretion. These studies demonstrate that net oxalate secretion occurs in the early portions of the proximal convoluted tubule, undergoes bidirectional transport of approximately equal magnitude in later segments of the proximal tubule, and probably is not transported in more distal nephron sites. The secretory mechanism for oxalate differs from that of PAH in that it is located in a different segment of the nephron and is not inhibited by probenecid. These differences suggest that the early portions of the proximal tubule are important in the renal metabolism of some organic acids.

Relationship between para-aminohippurate secretion and cellular morphology in rabbit proximal tubules.

Previous studies in the mammalian proximal tubule have suggested that para-aminohippurate (PAH) secretion is approximately threefold greater in the straight segment, or pars recta, than in the convoluted segment, or pars convoluta. However, the possibility that the site of maximal PAH secretion might be related better to particular tubule segments as identified by cell type had not been explored. In addition, the presence or absence of differences in PAH secretion between morphologically identical regions of superficial (SF) vs. juxtamedullary (JM) proximal tubules has not been examined. These issues were studied using a combination of histologic methods and measurement of [(3)H]PAH secretion in isolated perfused tubules. Measurements of microdissected SF and JM proximal tubules from young and adult rabbits revealed that SF proximal tubules were slightly but significantly longer than JM tubules ([young rabbits: SF, 8.69+/-SE 0.14 mm vs. JM, 7.97+/-SE 0.13 mm; P < 0.01] [adult rabbits: SF, 10.61+/-SE 0.28 mm; JM, 9.17+/-SE 0.19 mm; P < 0.001]). Light and electron microscopy revealed three sequential segments (S(1), S(2), and S(3)) along the length of SF and JM proximal tubules as defined by cell type. PAH secretion was measured in each of these three segments by the isolated perfused tubule technique. Net PAH secretion in fmol/mm per min in SF proximal tubules was: S(1), 281+/-SE 21; S(2), 1,508+/-SE 104; S(3), 318+/-SE 46. Corresponding values in JM proximal tubules were 353+/-SE 31, 1,391+/-SE 72, and 188+/-SE 23. Net PAH secretion did not differ between comparable segments of SF and JM proximal tubules. It is concluded that differences in PAH secretion along the proximal tubule correlate best with cell type rather than the arbitrary division of the proximal tubule into pars convoluta and pars recta according to its external configuration. Evidence of functional heterogeneity between comparable segments of SF and JM proximal tubules was not observed.

ostnatal development of renal function: micropuncture and clearance studies in the dog.

Postnatal renal development was studied in dogs between 2 and 77 days. Single, superficial nephrons were evaluated by micropuncture, concurrently with measurements of total renal function and morphometric analyses in the same animals. Glomerular filtration rate for the entire kidney increased linearly from 0.13 ml/min per g kidney weight at 2 days to 0.91 at 77 days. Extraction of p-aminohippurate increased from about 20 to 80%, and renal plasma flow per g kidney weight, measured as Cpah/Epah, increased threefold during the same period. Filtration fraction increased to the mature value during the first half of the postnatal period studied. The clearance of urea per unit of renal mass increased with age, whereas the fraction of filtered urea reabsorbed declined during the early part of the postnatal period. The pattern of fractional urea reabsorption may be due mainly to increased medullary recycling of urea and to a rise in the reabsorption of water from the medullary collecting duct. Urine osmolality was higher than plasma from birth onward and rose with age. Osmolal equality of collecting duct fluid and medullary interstitium reflected mature vasopressin (ADH)-induced water permeability. The rise in urinary concentration was predominantly due to increasing medullary sequestration of urea. Glomerular filtration rate of the superficial nephron increased from 3.2 nl/min at 21 days, when subcapsular nephrons were uniformly patent, to 23.1 at 77 days. Despite this rise in filtered load, fractional reabsorption of sodium and water in superficial proximal tubules was constant and at the mature level from the onset of intratubular perfusion. Changes in arterial plasma protein concentration, in filtration fraction, and in the hydrostatic pressure gradient between proximal tubule and peritubular capillary may interact to maintain glomerulotubular balance. The data, together with results of an accompanying morphological study, demonstrate a sequence of coordinated changes during postnatal renal maturation.