Live Imaging of Mitochondria in Kidney Tissue.

Kidneys are highly aerobic organs and their function is tightly coupled to mitochondrial energy production. Renal tubular cells, particularly the proximal tubule (PT), require an abundance of mitochondria to provide sufficient energy for regulating fluid and electrolyte balance. Meanwhile, mitochondrial defects are implicated in a range of different kidney diseases. Multiphoton microscopy (MP) is a powerful tool that allows detailed study of mitochondrial morphology, dynamics, and function in kidney tissue. Here, we describe how MP can be used to image mitochondria in kidney tubular cells, either ex vivo in tissue slices or in vivo in living rodents, using both endogenous and exogenous fluorescent molecules. Moreover, changes in mitochondrial signals can be followed in real time in response to different insults or stimuli, in parallel with other important readouts of kidney tubular function, such as solute uptake and trafficking.

Intravital imaging of real-time endogenous actin dysregulation in proximal and distal tubules at the onset of severe ischemia-reperfusion injury.

Severe renal ischemia-reperfusion injury (IRI) can lead to acute and chronic kidney dysfunction. Cytoskeletal modifications are among the main effects of this condition. The majority of studies that have contributed to the current understanding of IRI have relied on histological analyses using exogenous probes after the fact. Here we report the successful real-time visualization of actin cytoskeletal alterations in live proximal and distal tubules that arise at the onset of severe IRI. To achieve this, we induced fluorescent actin expression in these segments in rats with hydrodynamic gene delivery (HGD). Using intravital two-photon microscopy we then tracked and quantified endogenous actin dysregulation that occurred by subjecting these animals to 60 min of bilateral renal ischemia. Rapid (by 1-h post-reperfusion) and significant (up to 50%) declines in actin content were observed. The decline in fluorescence within proximal tubules was significantly greater than that observed in distal tubules. Actin-based fluorescence was not recovered during the measurement period extending 24 h post-reperfusion. Such injury decimated the renal architecture, in particular, actin brush borders, and hampered the reabsorptive and filtrative capacities of these tubular compartments. Thus, for the first time, we show that the combination of HGD and intravital microscopy can serve as an experimental tool to better understand how IRI modifies the cytoskeleton in vivo and provide an extension to current histopathological techniques.

Immunotoxin SS1P is rapidly removed by proximal tubule cells of kidney, whose damage contributes to albumin loss in urine.

Recombinant immunotoxins (RITs) are chimeric proteins composed of an Fv and a protein toxin being developed for cancer treatment. The Fv brings the toxin to the cancer cell, but most of the RITs do not reach the tumor and are removed by other organs. To identify cells responsible for RIT removal, and the pathway by which RITs reach these cells, we studied SS1P, a 63-kDa RIT that targets mesothelin-expressing tumors and has a short serum half-life. The major organs that remove RIT were identified by live mouse imaging of RIT labeled with FNIR-Z-759. Cells responsible for SS1P removal were identified by immunohistochemistry and intravital two-photon microscopy of kidneys of rats. The primary organ of SS1P removal is kidney followed by liver. In the kidney, SS1P passes through the glomerulus, is taken up by proximal tubular cells, and transferred to lysosomes. In the liver, macrophages are involved in removal. The short half-life of SS1P is due to its very rapid filtration by the kidney followed by degradation in proximal tubular cells of the kidney. In mice treated with SS1P, proximal tubular cells are damaged and albumin in the urine is increased. SS1P uptake by kidney is reduced by coadministration of l-lysine. Our data suggests that l-lysine administration to humans might prevent SS1P-mediated kidney damage, reduce albumin loss in urine, and alleviate capillary leak syndrome.

Transport mechanism and affinity of [99mTc]Tc-mercaptoacetyltriglycine ([99mTc]MAG3) on the apical membrane of renal proximal tubule cells.

Technetium-99m-labeled mercaptoacetyltriglycine ([99mTc]MAG3) is widely used for evaluation of transplanted kidneys, diagnosis of tubular necrosis, and scintigraphic studies of tubular function. [99mTc]MAG3 is a substrate for organic anion transporter (OAT)1 and OAT3 on the basolateral membrane side for renal secretion. We investigated the transport mechanism and affinity of [99mTc]MAG3 on the apical membrane of renal proximal tubule cells for renal secretion. Adenosine triphosphate-binding cassette (ABC) transporters for renal secretion of [99mTc]MAG3 were examined using ABC transporter vesicles expressing multiple drug resistance 1 (MDR1), breast cancer resistance protein (BCRP), multidrug resistance-associated protein (MRP)2, and MRP4. MK-571, a MRP inhibitor, was applied to measure the Km and Vmax of MRP2 and MRP4 in a vesicle transport assay. Single photon emission computed tomography (SPECT) was performed in normal rats and MRP2-deficient Eisai hyperbilirubinuria rats (EHBR) using [99mTc]MAG3 with and without MK-571. [99mTc]MAG3 uptake in adenosine triphosphate was significantly higher than that in adenosine monophosphate in vesicles that highly expressed MRP2 and MRP4. The affinity of [99mTc]MAG3 for MRP4 was higher than that for MRP2. Renal secretion via MRP2 and MRP4 was identified by comparing normal and EHBR rats with and without MK-571 on SPECT. [99mTc]MAG3 is transported via MRP2 and MRP4 on the apical membrane of renal proximal tubule cells. The affinity of MRP4 is higher than that of MRP2. SIGNIFICANCE STATEMENT: [99mTc]MAG3, widely used for evaluation of transplanted kidneys, diagnosis of tubular necrosis, and scintigraphic studies of tubular function, is transported via MRP2 and MRP4 on the apical membrane of renal proximal tubule cells. The affinity of MRP4 is higher than that of MRP2.

Heterozygous Pkhd1C642* mice develop cystic liver disease and proximal tubule ectasia that mimics radiographic signs of medullary sponge kidney.

Heterozygosity for human polycystic kidney and hepatic disease 1 ( PKHD1) mutations was recently associated with cystic liver disease and radiographic findings resembling medullary sponge kidney (MSK). However, the relevance of these associations has been tempered by a lack of cystic liver or renal disease in heterozygous mice carrying Pkhd1 gene trap or exon deletions. To determine whether heterozygosity for a smaller Pkhd1 defect can trigger cystic renal disease in mice, we generated and characterized mice with the predicted truncating Pkhd1C642* mutation in a region corresponding to the middle of exon 20 cluster of five truncating human mutations (between PKHD1G617fs and PKHD1G644*). Mouse heterozygotes or homozygotes for the Pkhd1C642* mutation did not have noticeable liver or renal abnormalities on magnetic resonance images during their first weeks of life. However, when aged to ~1.5 yr, the Pkhd1C642* heterozygotes developed prominent cystic liver changes; tissue analyses revealed biliary cysts and increased number of bile ducts without signs of congenital hepatic fibrosis-like portal field inflammation and fibrosis that was seen in Pkhd1C642* homozygotes. Interestingly, aged female Pkhd1C642* heterozygotes, as well as homozygotes, developed radiographic changes resembling MSK. However, these changes correspond to proximal tubule ectasia, not an MSK-associated collecting duct ectasia. In summary, by demonstrating that cystic liver and kidney abnormalities are triggered by heterozygosity for the Pkhd1C642* mutation, we provide important validation for relevant human association studies. Together, these investigations indicate that PKHD1 mutation heterozygosity (predicted frequency 1 in 70 individuals) is an important underlying cause of cystic liver disorders and MSK-like manifestations in a human population.

Combined Structural and Functional Imaging of the Kidney Reveals Major Axial Differences in Proximal Tubule Endocytosis.

BACKGROUND: The kidney proximal convoluted tubule (PCT) reabsorbs filtered macromolecules via receptor-mediated endocytosis (RME) or nonspecific fluid phase endocytosis (FPE); endocytosis is also an entry route for disease-causing toxins. PCT cells express the protein ligand receptor megalin and have a highly developed endolysosomal system (ELS). Two PCT segments (S1 and S2) display subtle differences in cellular ultrastructure; whether these translate into differences in endocytotic function has been unknown. METHODS: To investigate potential differences in endocytic function in S1 and S2, we quantified ELS protein expression in mouse kidney PCTs using real-time quantitative polymerase chain reaction and immunostaining. We also used multiphoton microscopy to visualize uptake of fluorescently labeled ligands in both living animals and tissue cleared using a modified CLARITY approach. RESULTS: Uptake of proteins by RME occurs almost exclusively in S1. In contrast, dextran uptake by FPE takes place in both S1 and S2, suggesting that RME and FPE are discrete processes. Expression of key ELS proteins, but not megalin, showed a bimodal distribution; levels were far higher in S1, where intracellular distribution was also more polarized. Tissue clearing permitted imaging of ligand uptake at single-organelle resolution in large sections of kidney cortex. Analysis of segmented tubules confirmed that, compared with protein uptake, dextran uptake occurred over a much greater length of the PCT, although individual PCTs show marked heterogeneity in solute uptake length and three-dimensional morphology. CONCLUSIONS: Striking axial differences in ligand uptake and ELS function exist along the PCT, independent of megalin expression. These differences have important implications for understanding topographic patterns of kidney diseases and the origins of proteinuria.

Selective Nanoparticle Targeting of the Renal Tubules.

Direct targeting to the kidneys is a promising strategy to improve drug therapeutic index for the treatment of kidney diseases. We sought to investigate the renal selectivity and safety of kidney-targeted mesoscale nanoparticle technology. We found that direct intravenous administration of these particles resulted in 26-fold renal selectivity and localized negligibly in the liver or other organs. The nanoparticles targeted the renal proximal tubular epithelial cells, as evidenced by intravital microscopy and ex vivo imaging. Mice treated with the nanoparticles exhibited no negative systemic consequences, immune reaction, liver impairment, or renal impairment. The localization of material selectively to the renal tubules is uncommon, and this work portends the development of renal-targeted drugs for the treatment of kidney diseases.

Outcome of renal proximal tubular dysfunction with Fanconi syndrome caused by sodium valproate.

BACKGROUND: Although Fanconi syndrome is rare in patients with epilepsy treated with sodium valproate (VPA), the prevalence might be higher in children with severe motor and intellectual disabilities (SMID). VPA-induced Fanconi syndrome usually has a favorable outcome, but the long-term outcome of renal tubular dysfunction in SMID patients remains unknown. The aim of this study was therefore to investigate the long-term outcome of renal proximal dysfunction in SMID children with Fanconi syndrome caused by VPA. METHODS: The records of six children with SMID and Fanconi syndrome caused by VPA were retrospectively reviewed to assess long-term proximal renal tubular function after discontinuation of VPA. All six patients had intractable epilepsy and required tube feeding. RESULTS: Proximal tubular dysfunction improved in almost all patients after VPA discontinuation, although abnormal uric acid reabsorption persisted in three patients. Five patients had hypocarnitinemia. After carnitine supplementation, one of these three patients with decreased ability to reabsorb uric acid had a normal serum level and improved fractional excretion of uric acid. CONCLUSIONS: Secondary carnitine deficiency may cause prolonged tubular dysfunction in some SMID patients with VPA-induced Fanconi syndrome. Fanconi syndrome caused by VPA is a usually reversible dysfunction of the proximal tubules, but can be permanent. Although not effective for all patients, carnitine is recommended for patients with VPA-induced Fanconi syndrome, especially children with SMID.

In vivo quantification of renal function in mice using clinical gamma cameras.

INTRODUCTION: In preclinical research, the growing number of transgenic models has led to the need for renal-function studies in mice. Many efforts have been made to develop dedicated SPECT systems for rodents, but their availability is limited due to high capital costs. The aim of this work is to demonstrate the feasibility of mouse renal imaging by using an inexpensive alternative based on clinical gamma-cameras. METHODS: A healthy mouse was scanned 3 h after injection of 6 mCi of Dimercaptosuccinic acid (DMSA) labeled with 99mTc by using a single-head gamma-camera in conjunction with a dedicated pinhole collimator. List-mode data were binned to emulate multiple injections of 1 mCi, 0.1 mCi and 0.01 mCi of 99mTc-DMSA and 6-min ventral and dorsal planar images were acquired and SPECT imaging (60 projection images acquired over 60 min) was performed. An optimization of the protocols in terms of injected activity, time scan, renal cortex uniformity and cortex-to-pelvis contrast was carried out. RESULTS: The appropriate protocols were an injected activity of 0.6 mCi, combined with duration of scanning of 1 min for planar and 60 min for SPECT imaging. Our results were validated through the relative quantification of renal function, which showed that both kidneys contributed equally to the total function. They showed that functional structures of the mouse kidneys can be visually distinguished as easily as in human studies. CONCLUSIONS: Our findings showed the feasibility of conducting quantitative DMSA SPECT studies of anesthetized mice on clinical gamma cameras.

Single photon emission-computed tomography (SPECT) for functional investigation of the proximal tubule in conscious mice.

Noninvasive analysis of renal function in conscious mice is necessary to optimize the use of mouse models. In this study, we evaluated whether single photon emission-computed tomography (SPECT) using specific radionuclear tracers can be used to analyze changes in renal proximal tubule functions. The tracers included (99m)TC- dimercaptosuccinic acid ((99m)Tc-DMSA), which is used for cortex imaging; (99m)Tc-mercaptoacetyltriglycine ((99m)Tc-MAG3), used for dynamic renography; and (123)I-beta(2)-microglobulin, which monitors receptor-mediated endocytosis. (99m)Tc-DMSA SPECT imaging was shown to delineate the functional renal cortex with a approximately 1-mm spatial resolution and accumulated in the cortex reaching a plateau 5 h after injection. The cortical uptake of (99m)Tc-DMSA was abolished in Clcn5 knockout mice, a model of proximal tubule dysfunction. Dynamic renography with (99m)Tc-MAG3 in conscious mice demonstrated rapid extraction from blood, renal accumulation, and subsequent tubular secretion. Anesthesia induced a significant delay in the (99m)Tc-MAG3 clearance. The tubular reabsorption of (123)I-beta(2)-microglobulin was strongly impaired in the Clcn5 knockout mice, with defective tubular processing and loss of the native tracer in urine, reflecting proximal tubule dysfunction. Longitudinal studies in a model of cisplatin-induced acute tubular injury revealed a correlation between tubular recovery and (123)I-beta(2)-microglobulin uptake. These data show that SPECT imaging with well-validated radiotracers allows in vivo investigations of specific proximal tubule functions in conscious mice.

Visualization of three-dimensional nephron structure with microcomputed tomography.

The three-dimensional architecture of nephrons in situ and their interrelationship with other nephrons are difficult to visualize by microscopic methods. The present study uses microcomputed X-ray tomography (micro-CT) to visualize intact nephrons in situ. Rat kidneys were perfusion-fixed with buffered formalin and their vasculature was subsequently perfused with radiopaque silicone. Cortical tissue was stained en bloc with osmium tetroxide, embedded in plastic, scanned, and reconstructed at voxel resolutions of 6, 2, and 1 microm. At 6 microm resolution, large blood vessels and glomeruli could be visualized but nephrons and their lumens were small and difficult to visualize. Optimal images were obtained using a synchrotron radiation source at 2 microm resolution where nephron components could be identified, correlated with histological sections, and traced. Proximal tubules had large diameters and opaque walls, whereas distal tubules, connecting tubules, and collecting ducts had smaller diameters and less opaque walls. Blood vessels could be distinguished from nephrons by the luminal presence of radiopaque silicone. Proximal tubules were three times longer than distal tubules. Proximal and distal tubules were tightly coiled in the outer cortex but were loosely coiled in the middle and inner cortex. The connecting tubules had the narrowest diameters of the tubules and converged to form arcades that paralleled the radial vessels as they extended to the outer cortex. These results illustrate a potential use of micro-CT to obtain three-dimensional information about nephron architecture and nephron interrelationships, which could be useful in evaluating experimental tubular hypertrophy, atrophy, and necrosis.

Renal tubular dysgenesis: report of two cases in a non-consanguineous couple and review of the literature.

Renal tubular dysgenesis is a rare congenital abnormality of renal development characterized by short and poorly developed proximal convoluted tubules. It is associated to late-appearing oligohydramnios, Potter's sequence, pulmonary hypoplasia and calvarial bone hypoplasia with enlarged fontanels. The onset of oligohydramnios is delayed and variable, normally later than 22 weeks of gestation, conditioning variation in the expression of the other deformities. We report 2 cases of renal tubular dysgenesis occurring in a non-consanguineous couple.

Effect of nitric oxide on ammoniagenesis in rats.

AIM: This in vitro study using rat cortical slices, isolated proximal tubules and mitochondria was conducted to investigate the effect of exogenous and endogenous nitric oxide on ammoniagenesis. METHODS AND RESULTS: The cortical slices were incubated with phosphate-buffered saline containing 1 mML-glutamine at 37 degrees C andglutamine-stimulated ammoniagenesis which was further elevated with 10(-7)M ANGII showed a time-dependent decrease during 2 h. 10(-4)M L-NAME or 10(-5)ML-canavanin caused a similar ammonia elevation to that of ANGII, whereas the addition of 10(-5)M SNAP attenuated the ammonia-increasing effects of ANGII and L-NAME. Basal or exogenous NO without significantly affecting glutamine uptake of the slices seemed to convert the glutamine deamidation pathway to transamination, since L-NAME increased the ammonia to glutamine ratio from 0.87 +/- 0.08 mol/mol to 1.03 +/- 0.04 (p < 0.01). L-NAME increased both ammoniagenesis and mitochondrial oxygen consumption but SNAP depressed them. Endogenous NO reduced ammoniagenesis without changing the mitochondrial permeability transition pore (PTP), whereas exogenous NO-induced attenuation in ammoniagenesis was associated with elevated PTP in a CsA-sensitive manner. CONCLUSION: These results demonstrated that in rat kidney, basal NO depresses mitochondrial oxygen consumption and attenuates ammoniagenesis without affecting PTP; however, exogenous NO inhibits ammonia production by disturbing PTP in isolated mitochondria.

Megalin is essential for renal proximal tubule reabsorption of (111)In-DTPA-octreotide.

UNLABELLED: Radiolabeled somatostatin analogs have been shown to be important radiopharmaceuticals for tumor diagnosis and radionuclide therapy. The kidney has appeared to be the critical organ during radionuclide therapy because of peptide reabsorption and retention in the proximal tubules after glomerular filtration. The molecular mechanism of renal reabsorption of these analogs has not been clarified. A possible receptor candidate is megalin, a multiligand scavenger receptor in the renal proximal tubules. The objective of this study was to investigate the role of megalin in tubular reabsorption of radiolabeled somatostatin analogs by using kidney-specific megalin-deficient mice versus mice with normal renal megalin expression. [(111)In-Diethylenetriaminepentaacetic acid (DTPA)]octreotide was used as a practical model of peptide. METHODS: Renal uptake of [(111)In-DTPA]octreotide was determined by animal SPECT scintigraphy at different time points after injection of the tracer and by measurement of radioactivity after isolation of the organs. Furthermore, ex vivo autoradiography of renal sections revealed the zonal distribution of radioactivity in the megalin-deficient and megalin-expressing kidneys. RESULTS: SPECT scintigraphy of [(111)In-DTPA]octreotide at 3 and 24 h after injection clearly showed lower renal radioactivity in megalin-deficient kidneys than in megalin-expressing kidneys, both in male and in female mice, in accordance with counts obtained after isolation of the organ (70%-85% reduction of uptake in the megalin-deficient kidneys, P < 0.001). Renal uptake of [(111)In-DTPA]octreotide was significantly higher in female than in male kidneys (P < 0.001). Ex vivo autoradiograms clearly showed that renal radioactivity was not homogeneously distributed in the megalin-expressing kidneys but localized in the renal cortex. Quantification of the autoradiogram data confirmed the reduced radioactivity in the renal cortex of megalin-deficient kidneys. CONCLUSION: This study revealed the molecular mechanism of [(111)In-DTPA]octreotide uptake in renal proximal tubules involving the receptor megalin. Identification of megalin may be crucial for further research into strategies to reduce renal uptake.

How to interpret a deterioration of split function?

OBJECTIVE: A drop of split renal function often constitutes a criterion for pyeloplasty in hydronephrosis since it is considered as representing deterioration of the affected kidney. The aim of this work was to determine, in a selected population of patients with a drop of split function of at least 5%, if the evolution of single kidney glomerular filtration rate (SKGFR) was parallel to the one of split renal function. METHODS: From a large data basis, we found retrospectively only 29 children (10 below and 19 above two years of age at first examination) having had at least two Tc-99m mertiatide (Tc-99m MAG3) renographic explorations for various urological diseases, with a decrease of split function of at least 5% between the two examinations. Evolution of split function was compared to evolution of SKGFR obtained by means of the combination of Tc-99m MAG3 split function and overall glomerular filtration rate as given by the chromium Cr 51 ethylenediamine tetraacetic acid (EDTA) clearance. RESULTS: For the group above two years of age, SKGFR increased or remained stable in 63% of the cases, while in the children less than 2 years of age, a decrease of SKGFR was never observed, according to the maturation of overall GFR in this age group. Thus, the decrease of split function was not necessarily associated with a similar decrease of SKGFR. CONCLUSION: In patients with unilateral or bilateral urological disorders, deterioration of split renal function does not necessary correspond to a loss of function of the affected kidney. SKGFR often modifies the interpretation of split function.

Role of connective tissue growth factor in mediating hypertrophy of human proximal tubular cells induced by angiotensin II.

BACKGROUND/AIMS: Cellular hypertrophy is an early important pathological feature of renal diseases such as diabetic nephropathy and remnant kidney. Recent studies have demonstrated that angiotensin II (Ang II) might be a major contributor in regulating cell hypertrophy. However, the exact mechanism is still unclear. The aim of our present work was to investigate the possible role of a newly clarified fibrogenic factor, connective tissue growth factor (CTGF), in mediating the effect of Ang II-induced tubular cell hypertrophy. METHODS: The cell line HK2 was grown in Dulbecco's modified Eagle's medium containing 10% heat-inactivated fetal calf serum. After the cells were rested in serum-free medium for 24 h, the dose and time response of CTGF mRNA expression to Ang II stimulation was observed by RT-PCR, and protein synthesis was observed by Western blotting. The effect of anti-CTGF on Ang II (10(-7)M)-induced [(3)H]-leucine incorporation, total protein content (Coomassie brilliant blue G250 method) and change in cellular size [determined by a scanning electronic microscope (SEM)] was also observed. The influence of anti-CTGF antibody on the cell cycle was analyzed by using a fluorescence-activated cell sorter flow cytometer. RESULTS: The results showed that Ang II induced expression of CTGF mRNA in a time- and dose-dependent manner (p < 0.05 and 0.01, respectively). Stimulation of cells with Ang II (10(-7)M) for 48 h resulted in a 92% increase in [(3)H]-leucine incorporation (5,584 cpm/10(5) cells at 0 h vs. 10,741 cpm/10(5) cells at 48 h; p = 0.01), which was significantly abolished by treatment with anti-CTGF antibody. Ang II (10(-7)M) significantly increased the total protein content in HK2 cells (control: 0.169 +/- 0.011 mg/10(5) cells vs. Ang II: 0.202 +/- 0.010 mg/10(5) cells; p = 0.03), which was markedly inhibited by cotreatment with anti-CTGF antibody. The average cellular diameter determined by SEM showed that the increase in cell size induced by Ang II could be significantly inhibited by anti-CTGF antibody (control: 11.92 +/- 1.62 microm, Ang II group: 20.63 +/- 3.83 microm, Ang II + anti-CTGF group: 16.43 +/- 3.23 microm; p < 0.01, respectively). Furthermore, the flow cytometer study showed that Ang II arrested the cell cycle at G0-G1 phase, which was significantly reversed by treatment with anti-CTGF antibody (G0-G1 percentage: in Ang II group: 76 +/- 1.8%, in Ang II + anti-CTGF group: 71 +/- 1.78%; p = 0.04). CONCLUSION: Our data are the first to clearly demonstrate that CTGF might be an important mediator of Ang II-induced renal hypertrophy, which suggests that inhibiting the production of CTGF might be the new strategy in early prevention of renal fibrosis.

Assessment of regional kidney function may provide new clinical understanding and assist in treatment of children with prenatal hydronephrosis.

PURPOSE: As calculated from a (99m)technetium (Tc)-mercaptoacetyltriglycine (MAG3) renogram, differential renal function is an important parameter affecting the clinical treatment of children with prenatal unilateral hydronephrosis. We determined whether value is potentially added by calculating a functional image from MAG3 renograms that represents proximal tubule ability to clear tracer from the blood, that is the MAG3 clearance image. MATERIALS AND METHODS: MAG3 clearance image findings and differential renal function at presentation and followup in 59 nonsurgically and 42 surgically treated children with prenatally diagnosed unilateral hydronephrosis were retrospectively reviewed. All patients underwent at least 3 99mTc-MAG3 renograms. RESULTS: In the surgical and nonsurgical groups there was no significant difference in followup (p = 0.11), age at presentation (p = 0.98) or age at last visit (p = 0.97), whereas differential renal function was significantly different (p = 0.01). A large affected kidney with focal defects was the most frequent finding in each group, including kidneys with a normal differential renal function of 45% to 55%. In the nonsurgical group the most common observation was improvement in the MAG3 clearance image in 26 of 59 cases (44%) and stabilized differential renal function in 41 (70%). Postoperatively the MAG3 clearance image improved in 31 of 42 cases (74%) and differential renal function remained stable in 25 (60%), while no significant increase in differential renal function was noted at the last visit (p = 0.74). CONCLUSIONS: This study shows that it is possible to assess regional parenchymal function using the MAG3 clearance image in children with unilateral hydronephrosis. This information is available in addition to 99mTc-MAG3 information on the whole kidney, that is differential renal function. The majority of affected kidneys with normal differential renal function showed parenchymal defects, suggesting regional renal dysfunction. However, functional improvement on a regional basis occurred in each group of children during followup.

Biphasic effects of dopamine on 86rubidium uptake in rat renal proximal tubules.

The mechanism(s) by which dopamine inhibits Na+-K+-ATPase activity in the renal proximal tubule is still controversial. We studied the short-term effects of dopamine on the sodium pump in rat renal proximal tubule suspensions with the 86Rb uptake method. Dopamine and the D1-like agonist, SKF81297, initially stimulated Na+-K+-ATPase activity at 5 min and subsequently inhibited it at 10 min and 20 min; the inhibition by 10 microM dopamine at 20 min was 21.3 +/- 4.5%. The inhibitory effect of dopamine on Na+-K+-ATPase activity was mimicked by thymeleatoxin (a classical protein kinase C [PKC] agonist) while Sp-8-CPT-cAMPS (a protein kinase A [PKA] agonist) had no effect. However, the combination of the PKC and PKA agonists mimicked the biphasic effects of dopamine and SKF81297. Rp-8-CPT-cAMPS (a PKA inhibitor), U-73122 (a phospholipase C inhibitor), or calphostin C (a PKC inhibitor), blocked the dopamine-mediated biphasic effects on Na+-K+-ATPase activity. It is suggested that the biphasic effects of dopamine on Na+-K+-ATPase activity (an initial stimulation and a subsequent inhibition) are transduced by activating both PKA and PKC through a D1-like receptor.

Poor renal accumulation of 99mTc-DMSA in idiopathic tubular proteinuria.

In Japanese patients idiopathic tubular proteinuria presents mainly as asymptomatic tubular low molecular weight proteinuria. This disease has recently been shown to resemble Dent's disease which is characterized by tubular proteinuria, hypercalciuria, rickets and eventual renal failure. We report on 4 children with idiopathic tubular proteinuria. Although they had normal renal function, as evidenced by serum creatinine or creatinine clearance, they had very poor renal accumulation of 99mTc-DMSA and the presence of large amounts of tracer in the bladder. Additionally, the patient with the largest amounts of tubular proteinuria had the poorest renal accumulation of the 4 patients. The renal accumulation of tracer decreased with time from a maximum at 10 min after injection. These findings demonstrate that the tracer, once taken to be confined to the proximal tubular cells, is immediately excreted to the tubular lumen. We suggest that poor renal accumulation of 99mTc-DMSA is very important in elucidating the mechanism of idiopathic tubular proteinuria, and that 99mTc-DMSA renoscintigraphy is useful in the evaluation of the patient's renal function over time.