Papillary Interstitium Research Articles

Multimodal imaging reveals a unique autofluorescence signature of Randall's plaque.

Kidney stones frequently develop as an overgrowth on Randall's plaque (RP) which is formed in the papillary interstitium. The organic composition of RP is distinct from stone matrix in that RP contains fibrillar collagen; RP in tissue has also been shown to have two proteins that are also found in stones, but otherwise the molecular constituents of RP are unstudied. We hypothesized that RP contains unique organic molecules that can be differentiated from the stone overgrowth by fluorescence. To test this, we used micro-CT-guided polishing to expose the interior of kidney stones for multimodal imaging with multiphoton, confocal and infrared microscopy. We detected a blue autofluorescence signature unique to RP, the specificity of which was also confirmed in papillary tissue from patients with stone disease. High-resolution mineral mapping of the stone also showed a transition from the apatite within RP to the calcium oxalate in the overgrowth, demonstrating the molecular and spatial transition from the tissue to the urine. This work provides a systematic and practical approach to uncover specific fluorescence signatures which correlate with mineral type, verifies previous observations regarding mineral overgrowth onto RP and identifies a novel autofluorescence signature of RP demonstrating RP's unique molecular composition.

Epac1 null mice have nephrogenic diabetes insipidus with deficient corticopapillary osmotic gradient and weaker collecting duct tight junctions.

The cAMP-mediator Epac1 (RapGef3) has high renal expression. Preliminary observations revealed increased diuresis in Epac1-/- mice. We hypothesized that Epac1 could restrict diuresis by promoting transcellular collecting duct (CD) water and urea transport or by stabilizing CD paracellular junctions to reduce osmolyte loss from the renal papillary interstitium. In Epac1-/- and Wt C57BL/6J mice, renal papillae, dissected from snap-frozen kidneys, were assayed for the content of key osmolytes. Cell junctions were analysed by transmission electron microscopy. Urea transport integrity was evaluated by urea loading with 40% protein diet, endogenous vasopressin production was manipulated by intragastric water loading and moderate dehydration and vasopressin type 2 receptors were stimulated selectively by i.p.-injected desmopressin (dDAVP). Glomerular filtration rate (GFR) was estimated as [14 C]inulin clearance. The glomerular filtration barrier was evaluated by urinary albumin excretion and microvascular leakage by the renal content of time-spaced intravenously injected 125 I- and 131 I-labelled albumin. Epac1-/- mice had increased diuresis and increased free water clearance under antidiuretic conditions. They had shorter and less dense CD tight junction (TJs) and attenuated corticomedullary osmotic gradient. Epac1-/- mice had no increased protein diet-induced urea-dependent osmotic diuresis, and expressed Wt levels of aquaporin-2 (AQP-2) and urea transporter A1/3 (UT-A1/3). Epac1-/- mice had no urinary albumin leakage and unaltered renal microvascular albumin extravasation. Their GFR was moderately increased, unless when treated with furosemide. Our results conform to the hypothesis that Epac1-dependent mechanisms protect against diabetes insipidus by maintaining renal papillary osmolarity and the integrity of CD TJs.

MP11-04 NEW INSIGHTS INTO THE GENESIS AND ADHERENCE OF URINARY CALCULI USING HIGH RESOLUTION MICROSCOPY

You have accessJournal of UrologyImaging/Radiology: Uroradiology II1 Apr 2015MP11-04 NEW INSIGHTS INTO THE GENESIS AND ADHERENCE OF URINARY CALCULI USING HIGH RESOLUTION MICROSCOPY Sunita Ho, Frances Allen, Andrew Minor, Sabra Djomehri, Ling Chen, Thomas Chi, Krishna Ramaswamy, and Marshall Stoller Sunita HoSunita Ho More articles by this author , Frances AllenFrances Allen More articles by this author , Andrew MinorAndrew Minor More articles by this author , Sabra DjomehriSabra Djomehri More articles by this author , Ling ChenLing Chen More articles by this author , Thomas ChiThomas Chi More articles by this author , Krishna RamaswamyKrishna Ramaswamy More articles by this author , and Marshall StollerMarshall Stoller More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.385AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES It has been postulated that Randall's plaque (RP) is the nidus for calculus formation. What keeps the RP or the early “calculus” adherent to the papilla and allow it to grow? The purpose of this study is to provide insights by identifying mineral density variation and ultrastructure of the anchor between the calculus and parenchyma. METHODS 500 µm biopsies from non-stone forming and stone forming human papillae were obtained from patients and were immediately imaged using high resolution x-ray microscopy with a spatial resolution <1 µm. Post processing software was used to analyze tomograms to identify mineral density of the lumen, anchor and papillary interstitium. RESULTS A representative figure illustrates a 3-dimensional digital reconstruction (20x, far left) and virtual sections (40x, right two images) with a representative calculus emerging from a uriniferous tubule (yellow arrows). Multiple mineral gradients from higher to lower mineral densities (40X images) were observed. The mineral density of the approximately 200 µm formed “calculus” (1.3 g/cc, yellow arrow) was similar to that of the luminal wall (1.2 g/cc, white arrows). The 20–50 µm wide “anchor” (black arrow) attaching the formed “calculus” to the luminal wall illustrates similar heterogeneous mineral density (yellow arrows, 40X) as identified by a mixture of higher and lower x-ray attenuation regions. The rough surface morphology of the formed mineral/calculus (yellow arrow, 20X) was distinctly different than the mineralized structures within the luminal walls and the interstitium (red star) of the papilla (white arrows, 40X). CONCLUSIONS High resolution x-ray microscopy illustrates the anchor (attachment site) of urinary calculus emerging from the mineralized wall of a plaque-ridden uriniferous tubule. This anchor appears to be an integral part of the mineralized luminal wall. These results provide new insights and an appreciation for the genesis and attachment mechanism(s) of the calculus to the parenchyma. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e120 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Sunita Ho More articles by this author Frances Allen More articles by this author Andrew Minor More articles by this author Sabra Djomehri More articles by this author Ling Chen More articles by this author Thomas Chi More articles by this author Krishna Ramaswamy More articles by this author Marshall Stoller More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Sulfatides are required for renal adaptation to chronic metabolic acidosis

Urinary ammonium excretion by the kidney is essential for renal excretion of sufficient amounts of protons and to maintain stable blood pH. Ammonium secretion by the collecting duct epithelia accounts for the majority of urinary ammonium; it is driven by an interstitium-to-lumen NH3 gradient due to the accumulation of ammonium in the medullary and papillary interstitium. Here, we demonstrate that sulfatides, highly charged anionic glycosphingolipids, are important for maintaining high papillary ammonium concentration and increased urinary acid elimination during metabolic acidosis. We disrupted sulfatide synthesis by a genetic approach along the entire renal tubule. Renal sulfatide-deficient mice had lower urinary pH accompanied by lower ammonium excretion. Upon acid diet, they showed impaired ammonuria, decreased ammonium accumulation in the papilla, and chronic hyperchloremic metabolic acidosis. Expression levels of ammoniagenic enzymes and Na(+)-K(+)/NH4(+)-2Cl(-) cotransporter 2 were higher, and transepithelial NH3 transport, examined by in vitro microperfusion of cortical and outer medullary collecting ducts, was unaffected in mutant mice. We therefore suggest that sulfatides act as counterions for interstitial ammonium facilitating its retention in the papilla. This study points to a seminal role of sulfatides in renal ammonium handling, urinary acidification, and acid-base homeostasis.

Peginesatide Clearance, Distribution, Metabolism, and Excretion in Monkeys following Intravenous Administration

Peginesatide, a polyethylene glycol (PEG)ylated peptide-based erythropoiesis-stimulating agent, stimulates the erythropoietin receptor dimer that governs erythropoiesis. Studies were designed to determine the erythropoietic response, pharmacokinetics (PK), tissue distribution, metabolism, and excretion of peginesatide in nonhuman primates following a single i.v. dose. The PK profile of peginesatide (0.1-5 mg/kg) is characterized by low, dose-dependent plasma clearance; small volume of distribution; and long half-life. The peginesatide PK profile following a single i.v. dose is consistent with the sustained erythropoiesis. Biodistribution quantitative whole-body autoradiography demonstrated high peginesatide levels in bone marrow (i.e., primary hematopoietic site) as well as other known hematopoietic sites persisting through at least 3 weeks at 2.1 mg/kg. Microautoradiography analysis at 48 hours postdose revealed uniform and high distribution of radioactivity in the bone marrow and splenic red pulp with less extensive distribution in the renal cortex (glomeruli, associated ducts, interstitial cells). Radioactivity in the kidney was most prominent in the outer medullary and papillary interstitium. At 2 weeks after dosing, cumulative radioactivity recovery in the urine and feces was 60 and 7% of the administered dose, respectively, with most of the radioactivity associated with the parent molecule. In conclusion, the PK characteristics are consistent with a PEGylated peptide of a 45-kDa molecular mass, specifically low volume of distribution and long half-life. Drug was localized principally to hematopoietic sites, and nonspecific tissue retention was not observed. The nonhuman primate data indicate that peginesatide is metabolically stable and primarily excreted in the urine.

Urine Concentrating Mechanism: Impact of Vascular and Tubular Architecture and a Proposed Descending Limb Urea‐Na Cotransporter

We extended a region‐based mathematical model of the renal medulla of the rat kidney, previously developed by us, to represent new anatomic findings on the vascular architecture in the rat inner medulla (IM). The model represents separation of descending vasa recta from the descending limbs of loops of Henle, and a papillary segment of the descending thin limb that is water impermeable and highly urea permeable. Model results suggest that, despite compartmentalization of IM blood flow, IM interstitial fluid composition is substantially more homogeneous compared to the outer medulla (OM). We used the model to study medullary blood flow in antidiuresis and the effects of vascular countercurrent exchange. We also hypothesize that the terminal aquaporin‐1 null segment of the long descending thin limbs expresses a urea‐Na or urea‐Cl cotransporter. As urea diffuses from the urea‐rich papillary interstitium into the descending thin limb luminal fluid, NaCl is secreted via the cotransporter against its concentration gradient. That NaCl is then reabsorbed near the loop bend, raising the interstitial fluid osmolality and promoting water reabsorption from IM collecting ducts. The model predicts that the presence of the urea‐Na or urea‐Cl cotransporter facilitates cycling of NaCl within the IM, and yields a loop‐bend fluid composition consistent with experimental data.

Urine concentrating mechanism: impact of vascular and tubular architecture and a proposed descending limb urea-Na+cotransporter

We extended a region-based mathematical model of the renal medulla of the rat kidney, previously developed by us, to represent new anatomic findings on the vascular architecture in the rat inner medulla (IM). In the outer medulla (OM), tubules and vessels are organized around tightly packed vascular bundles; in the IM, the organization is centered around collecting duct clusters. In particular, the model represents the separation of descending vasa recta from the descending limbs of loops of Henle, and the model represents a papillary segment of the descending thin limb that is water impermeable and highly urea permeable. Model results suggest that, despite the compartmentalization of IM blood flow, IM interstitial fluid composition is substantially more homogeneous compared with OM. We used the model to study medullary blood flow in antidiuresis and the effects of vascular countercurrent exchange. We also hypothesize that the terminal aquaporin-1 null segment of the long descending thin limbs may express a urea-Na(+) or urea-Cl(-) cotransporter. As urea diffuses from the urea-rich papillary interstitium into the descending thin limb luminal fluid, NaCl is secreted via the cotransporter against its concentration gradient. That NaCl is then reabsorbed near the loop bend, raising the interstitial fluid osmolality and promoting water reabsorption from the IM collecting ducts. Indeed, the model predicts that the presence of the urea-Na(+) or urea- Cl(-) cotransporter facilitates the cycling of NaCl within the IM and yields a loop-bend fluid composition consistent with experimental data.

The role of hypoxia, increased oxygen consumption, and hypoxia-inducible factor-1 alpha in progression of chronic kidney disease

Tubulointerstitial hypoxia in the kidney has been considered a hallmark of injury and mediator of disease progression. This review focuses on hypoxia-inducible factor (HIF)-1, a master transcription factor in cellular adaptation to hypoxia. HIF-1alpha is expressed in the hypoxic tubular epithelium as well in as the papillary interstitium and glomerular epithelial cells. Although HIF-1 plays a protective role in a number of acute kidney injury models when overexpressed, its activation in chronic kidney disease results in multiple phenotypic changes, depending on the pathological context. Hypoxia, especially HIF-1, is a critical mediator in the pathogenesis of chronic kidney disease. Underlying molecular mechanisms, together with responsible HIF target genes, are currently under investigation.

Hyaluronan synthases and hyaluronidases in the kidney during changes in hydration status

Hyaluronan is a large glycosaminoglycan that is abundant in the interstitium of the renal medulla/papilla. Papillary hyaluronan increases during hydration and decreases during dehydration. Due to its gel properties and ability to retain large volumes of water, hyaluronan plays a role in renal water handling by affecting the permeability characteristics of the papillary interstitium. The focus of the present investigation was the regulation of hyaluronan metabolism in the kidney, especially during variations in hydration status.In control papillas, HAS 2 mRNA was heavily expressed and HAS 1 and 3 mRNA were weakly distributed. HYALs 1–3 mRNA were found at high expression and HYAL 4 was only weakly expressed. In hydrated animals, the diuretic response (12-fold) was followed by a 58% elevation in papillary hyaluronan and a 45% reduction in the excreted urinary hyaluronidase activity. No difference was determined in HAS 1–3 mRNA or HYAL 1, 3–4 mRNA expression, suggesting a change in activity rather than amount of protein. In dehydrated animals, antidiuresis was followed by a 22% reduction in papillary hyaluronan and a 62% elevation in excreted urinary hyaluronidase activity. Plasma vasopressin was 2.8-fold higher in dehydrated vs. hydrated rats.In conclusion, HAS 2 appears a major contributor to the baseline levels of hyaluronan. Reduced HAS 2 gene expression and increased excreted urinary hyaluronidase activity during dehydration contribute to the reduced amount of hyaluronan and to antidiuretic response.

Tubular and Interstitial Nephrocalcinosis

We determined whether nephrocalcinosis is common and whether its detection is influenced by renal tissue processing. Renal cortical and papillary tissue was obtained from the unaffected parts of 15 kidneys removed due to an oncological indication. The effect of tissue processing on the loss of crystals was studied in a kidney with nephrocalcinosis due to chronic pyelonephritis. Immediately frozen and formaldehyde fixed sections were analyzed by polarized light and Raman spectroscopy, and stained for calcium (Yasue) and hyaluronan. Although 13 of 15 snap-frozen sections from tumor kidneys contained birefringent particles (mean +/- SD 3.2 +/- 2.9 particles per cm(2)) in the renal tubules, this was not considered nephrocalcinosis because the crystals were not attached to the epithelial lining. Interstitial nephrocalcinosis was found on Yasue stain in 3 of 15 kidneys with tumor (20%). Calcium deposits were found in the papillary interstitium only, always together with hyaluronan. Formaldehyde fixed sections from the pyelonephritis kidney contained fewer renal tubular cell associated birefringent particles than immediately frozen sections (9.4 +/- 1.9 vs 41.6 +/- 1.2 per cm(2)). Particles were composed of calcium oxalate monohydrate (Yasue and Raman). There are 2 distinct forms of nephrocalcinosis, including tubular nephrocalcinosis, which seems to be reserved for specific conditions such as chronic pyelonephritis, and interstitial nephrocalcinosis. The incidence of tubular calcium oxalate nephrocalcinosis could be underestimated due to the loss of crystals during tissue processing for routine histology. The crystal binding molecule hyaluronan may have a role in the 2 forms of nephrocalcinosis.

A new animal model of hyperoxaluria and nephrolithiasis in rats with small bowel resection

An association between small bowel resection and stone disease has been noted, which is primarily due to increased gut oxalate absorption and resulting excretion by the kidney. In order to better understand the factors affecting both oxalate absorption and renal excretion, and the resulting renal lesions, we have developed a rodent model of small bowel resection and hyperoxaluria. Using this model, we have studied the renal histology in animals with hyperoxaluria over time spans from 2 weeks to 7 months. The initial lesion appears to be crystal formation along the brush border of the proximal tubule, with eventual crystal deposition in collecting ducts and papillary interstitium, and eventual tubule obstruction, interstitial inflammation and fibrosis. Crystal formation appears to dissociate from urinary supersaturation. We hypothesize that oxalate transporters in the proximal tubule may increase local saturations, leading to crystal formation at this site initially. Further studies are required to better characterize the causes and consequences of hyperoxaluria in this animal model.

Early Pathophysiological Features in Canine Renal Papillary Necrosis Induced by Nefiracetam

To ascertain the early pathophysiological features in canine renal papillary necrosis (RPN) caused by the neurotransmission enhancer nefiracetam, male beagle dogs were orally administered nefiracetam at 300 mg/kg/day for 4 to 7 weeks in comparison with ibuprofen, a non-steroidal anti-inflammatory drug (NSAID), at 50 mg/kg/day for 5 weeks. During the dosing period, the animals were periodically subjected to laboratory tests, light-microscopic, immunohistochemical, and electron-microscopic examinations and/or cyclooxygenase (COX)-2 mRNA analysis. In laboratory tests, a decrease in urinary osmotic pressure and increases in urine volume and urinary lactate dehydrogenase (LDH) level were early biomarkers for detecting RPN. Light-microscopically, nefiracetam revealed epithelial swelling and degeneration in the papillary ducts in week 7, while ibuprofen displayed degeneration and necrosis in the papillary interstitium in week 5. In immunohistochemical staining with COX-2 antibody, nefiracetam elicited a positive reaction within interstitial cells around the affected epithelial cells in the papillary ducts (upper papilla) in week 7, and ibuprofen positively reacted within interstitial cells adjacent to the degenerative and/or necrotic lesions in week 5. Ultrastructurally, nefiracetam exhibited reductions of intracellular interdigitation and infoldings of epithelial cells in the papillary ducts, whereas ibuprofen showed no changes in the identical portions. Thus, the early morphological change in the papilla brought about by nefiracetam was quite different from that elicited by ibuprofen. By the renal papillary COX-2 mRNA expression analysis, nefiracetam exceedingly decreased its expression in week 4, but markedly increased it in week 7, suggesting an induction of COX-2 mRNA by renal papillary lesions. These results demonstrate that the epithelial cell in the papillary ducts is the primary target site for the onset of RPN evoked by nefiracetam.

Nephrolithiasis and nephrocalcinosis in rats with small bowel resection

Intestinal resection (IR) may lead to hyperoxaluria and nephrolithiasis. A rat model of IR was developed, in which kidney stones form. We describe the urine chemistries and histopathologic features. Rats underwent resection of 40-45 cm of distal ileum (n=16) or sham resection (SR) (n=8), and were then fed a 1% Na oxalate, 0.02% Ca diet. After 1 week on the diet, 24 h urine samples were obtained for stone chemistries. At 4-7 months after surgery, kidneys were examined grossly and by light microscopy. The extent and location of crystallization was assessed by polarized light. Histochemistry and infrared spectroscopy were used to determine crystal composition. IR rats had higher urine oxalate excretion (P<0.01) and concentration (P<0.001) than SR rats, and lower urine citrate excretion; only IR rats formed kidney stones (12/15 surviving rats). Tissue calcification was found only in kidneys from IR rats, located in the cortex (83% of kidneys), medulla (73%) and papillary tip (47%). Crystals, composed of CaOx, apatite, and calcium carbonate, filled collecting duct lumens, and were associated with tubular obstruction, and interstitial inflammation. Crystals in the papillary interstitium incited inflammation with tubular destruction and development of progressive papillary erosion. This new rat model of nephrolithiasis and nephrocalcinosis resembles the pattern of urinary abnormalities and tissue calcification that may be seen in humans with small bowel resection. The model allows further studies of the mechanisms of renal crystal formation, and possible therapeutic interventions.

Drainage of plasma proteins from the renal medullary interstitium in rats.

1. Lymph vessels are scarce or lacking in the renal inner medulla, raising the question of whether plasma proteins entering the medullary interstitium are removed by diffusion through the interstitium to lymphatics in the outer medulla or cortex, or by convection into the vasa recta. 2. Using micropipettes, we infused 125I-albumin into the papilla of anaesthetized rats and watched its disappearance from the injection site as well as the uptake in the thoracic duct and plasma. 3. Tracer infused into the renal cortex appeared almost immediately in the thoracic duct lymph, and rose to a sevenfold higher concentration than in plasma, whereas tracer infused into the papilla appeared first and increased more sharply in plasma than in the lymph. No spread from the papillary injection site was observed. Tracer injected in renal hilar lymphatics was quantitatively recovered in the thoracic duct. 4. The plasma concentration pattern following papillary infusion was similar to that obtained by intravenous injection, indicating uptake in blood and subsequent distribution to extracellular fluid and lymph from all organs. 5. We conclude that plasma proteins normally diffusing out from the vasa recta are brought back through water flux (1) from the collecting ducts due to the high sodium chloride concentration in the papillary interstitium and (2) from the interstitium into the vasa recta driven by plasma protein osmotic pressure. Accordingly, there is no need for lymph vessels in the inner medulla.

High Resolution Radiography of Cadaveric Kidneys: Unraveling the Mystery of Randall's Plaque Formation

Purpose We used high resolution radiography to identify and characterize Randall's plaques in cadaveric kidneys. Materials and Methods A total of 50 consecutive sets of cadaveric kidneys was fixed, bivalved and imaged with micro-focal spot magnification radiography. Papillary calcifications were identified, localized and processed for light microscopy. Special immunohistochemical stains were implemented to aid localization of ectopic calcifications. Patient medical records and autopsy results were retrospectively evaluated and correlated with radiographic papillary calcifications. Results Of the 92 renal units with complete data 52 (57 percent) had radiographic evidence of renal medullary calcifications consistent with Randall's plaques. Unlike the original description of this condition, calcifications extended deep into the papilla. A history of hypertension was the only clinical parameter correlating with papillary calcifications. Calcium deposition was localized to the basement membrane of collecting tubules and vasa recta, and papillary interstitium. Conclusions Randall's plaques are not merely subepithelial deposits. Rather, they appear to extend deep within the papilla, and are intimately associated with collecting tubules and vasa recta. An association between papillary calcifications and urinary stone formation has yet to be proved but is under investigation.

Extravascular protein in the renal medulla: analysis by two methods

Two methods have been used to test for the presence of extravascular protein in the interstitium of the renal inner medulla. First, ascending vasa recta (AVR) segments were perfused with buffer containing 5 g/dl of albumin. The hydraulic pressure in the perfused vessel was varied to control transmembrane volume flux (Jv) to the interstitium. Interpolation to the point of zero Jv was employed to estimate interstitial Starling forces in the hydropenic rat papilla. Analysis of those experiments predicts that interstitial protein concentration (Ci) is high. When AVR segments are filled with oil, the oil column spontaneously breaks up as fluid is secreted into the lumen from the papillary interstitium. To obtain a lower limit on Ci, isolated AVR segments (IAS) filled with oil were sampled to measure protein concentration in the secreted fluid. In hydropenic rats, protein concentration was 3.4 +/- 0.5 and 5.2 +/- 0.2 g/dl in IAS and adjacent free-flowing AVR, respectively (P < 0.01). In rats subjected to furosemide and saline diuresis, the values were nearly identical, 4.7 +/- 0.2 and 5.2 +/- 0.2 g/dl, respectively. These separate experimental approaches corroborate a high concentration of protein in the renal inner medullary interstitium.

Molecular sieving of albumin by the ascending vasa recta wall.

Molecular sieving of albumin by ascending vasa recta. Evidence exists to support the presence of an extravascular pool of albumin in the renal medullary interstitium. This study used microperfusion in vivo to measure the transport of 125I-labeled albumin from descending (DVR) and ascending vasa recta (AVR) to the papillary interstitium. Perfusions were performed during furosemide diuresis with a buffer containing FITC-labeled dextran (FITC-Dx) 2 x 10(6) mol wt and 125I-albumin. Perfusate albumin and collection pressure were adjusted to induce either zero transcapillary volume flux (Jv) or high volume flux. When Jv was zero, the collectate-to-perfusate ratios of FITC-Dx (RDX) and 125I-albumin (Ralb) in the DVR and AVR were identical implying that diffusive efflux of albumin was immeasurably small. In contrast, when Jv was increased, paired comparison of Ralb and RDX in the same AVR revealed a difference, 1.58 +/- 0.06 vs 1.72 +/- 0.08, respectively (P less than 0.01). AVR perfusions in hydropenic animals showed similar results, Ralb = 1.70 +/- 0.07 and RDX = 2.00 +/- 0.07 (P less than 0.01). These data suggest that albumin transport across vasa recta in vivo is likely to be governed by solvent drag. The reflection coefficient of the AVR wall to 125I-albumin is estimated to be 0.78.

Resistance of ascending vasa recta to transport of water

A study was undertaken to determine the effect of increasing capillary pressure on volume flux in ascending vasa recta (AVR). In one experiment (group I), AVR were blocked by a single injection of paraffin wax and subjected to free-flow microperfusion at 10 nl/min. Collected fluid was obtained from the perfused vessels by micropuncture. In a second experiment (group II), AVR segments were isolated between two paraffin blocks and perfused at 10 nl/min. In group II, the collection pipette was pressurized to 0, 10, or 20 mmHg. Transmembrane volume flux was determined by measuring the change in concentration of fluorescein isothiocyanate-labeled dextran (2 x 10(6) mol wt) from perfusate to collected fluid. In group I, measurements revealed a capillary pressure of 10.3 +/- 0.5 (SE) mmHg and volume flux of 4.3 +/- 1.0 nl.mm-1.min-1. In group II, volume flux was 1.8 +/- 1.3, 5.9 +/- 1.0, and 11.2 +/- 1.1 nl.mm-1.min-1 at collection pressures of 0, 10, or 20 mmHg, respectively. Based on these data and an AVR diameter of 20 microns, AVR hydraulic conductivity is between 12.5 x 10(-6) and 18.7 x 10(-6) cm.s-1.mmHg-1. The papillary AVR have a high hydraulic conductivity. This is consistent with their role as the sole conduit for removal of water from the papillary interstitium.

Effect of sodium chloride gradients on water flux in rat descending vasa recta.

In the hydropenic kidney, volume efflux from descending vasa recta (DVR) occurs despite an intracapillary oncotic pressure that exceeds hydraulic pressure. That finding has been attributed to small solute gradients which may provide an additional osmotic driving force favoring water transport from DVR plasma to the papillary interstitium. To test this hypothesis, axial gradients of NaCl and urea in the papilla were eliminated by administration of furosemide and saline. DVR were then blocked with paraffin and microperfused at 10 nl/min with a buffer containing albumin, fluorescein isothiocyanate labeled dextran (FITC-Dx), 22Na, and NaCl in a concentration of 0 (hypotonic to the interstitium), 161 (isotonic) or 322 mM (hypertonic). Collectate was obtained from the perfused DVR by micropuncture and the collectate-to-perfusate ratios of FITC-Dx and 22Na were measured. A mathematical model was employed to determine DVR permeability (Ps) and reflection coefficient to NaCl (sigma NaCl). The rate of transport of water from the DVR lumen to the papillary interstitium was 2.8 +/- 0.3 (Nv = 22), -0.19 +/- 0.4 (Nv = 15), and -2.3 +/- 0.3 nl/min (Nv = 21) (mean +/- SE) when perfusate NaCl was 0, 161, or 322 mM, respectively (Nv = number of DVR perfused). The collectate-to-perfusate 22Na concentration ratios were 0.34 +/- 0.04, 0.36 +/- 0.04 and 0.37 +/- 0.03 for those groups, respectively. Based on these data, Ps is calculated to be 60.4 x 10(-5) +/- 4.0 x 10(-5) cm/s and sigma NaCl less than 0.05. The results of this study confirm that transcapillary NaCl concentrations gradients induce water movement across the wall of the DVR.

Glycosaminoglycans of the rat renomedullary interstitium: Ultrastructural and biochemical observations

The rat renal papillary interstitium which contains abundant proteoglycans is a unique area important in renal function. These proteoglycans were studied ultrastructurally by ruthenium red fixation and staining and phosphate-buffered fixation before and after enzyme digestion. A tissue culture of rat renomedullary interstitial cells, the predominant cell of the renal papillary interstitium, was studied for its ability to synthesize proteoglycans and the proteoglycans were then analyzed. Tissue slices of whole rat renal inner medulla were also evaluated for their synthetic ability. In combination, these studies indicate that the dominant glycosaminoglycan is hyaluronic acid. The tissue culture of rat renal medullary interstitial cells synthesized glycosaminoglycans and on analysis, hyaluronic acid was found to be the cheif glycosaminoglycan secreted by the renomedullary interstitial cells. Combined with the removal of the proteoglycans from tissue by leech hyaluronidase and testicular hyaluronidase, this suggests that the dominant glycosaminoglycan is hyaluronic acid. Hyaluronic acid is also synthesized by the intact papilla confirming the findings with the tissue culture. However, in addition, sulfated glycosaminoglycans were also synthesized by the intact papilla, presumably the product of the noninterstitial components of the papilla.