Search PubMed⌕ Search

Biomedical subjects

I Kurtz

Publications and source records attributed to I Kurtz.

At least 55 records · Page 3Linked to original sources

A new noninvasive quantification of renal blood flow with N-13 ammonia, dynamic positron emission tomography, and a two-compartment model.

In order to determine if dynamic positron emission tomography (PET) and N-13 ammonia can be used to quantitate regional RBF (rRBF) noninvasively, six anesthetized dogs were examined with PET imaging after an iv bolus administration of 5 mCi of N-13 ammonia. Renal time activity curves and the arterial input function were derived from regions of interest drawn over the renal cortex and abdominal aorta, respectively. For calculation of rRBF, less than 120 s of the initial data were used to minimize contamination by plasma metabolites of N-13 radioactivity. rRBF was quantitated with a two-compartment model, and the results were compared with simultaneously acquired microsphere blood flow measurement. Fourteen experiments were performed in six dogs, and four regions of interest on renal cortex were selected on each PET image. RBF derived from dynamic PET imaging with N-13 ammonia was linearly related to microsphere (MS) values (rRBF = 1.06 x MS - 0.17; r = 0.91). Mean rRBF in the canine experiments was 4.0 mL/min/g. The results indicate that dynamic N-13 ammonia renal PET can provide noninvasively quantitative rRBF.

Ammonia↗

Benzoate modulates renal and extrarenal nitrogen flow: metabolic mechanisms.

The mechanism by which benzoate enhances total nitrogen excretion was investigated in-situ and in separated rat renal proximal tubules. Orally administered benzoate augmented NH4+, urea and hippurate excretion 2, 1.9 and 76 fold respectively, as compared to baseline for control. Hippurate had similar effects. Benzoate augmented renal blood flow, glutamine extraction and total NH4+ production. Arterio-venous concentration differences of glutamine, glutamate, and NH4+ across the kidney, liver and gut demonstrated an increase in glutamine uptake by the kidney despite reduced release and uptake by the liver and gut, respectively; glutamate release by the kidney and gut was increased; NH4+ handling was unchanged at these three organs. Studies in separated rat renal proximal tubules demonstrated that benzoate stimulated glutamine dependent ammonia-genesis by activation of gamma-glutamyltransferase, via the synthesis of hippurate. The results demonstrate that benzoate can modulate the interorgan partitioning of nitrogen metabolites across several organs, the net effect of which is physiologically expressed as enhanced NH4+ , urea and hippurate excretion.

Ammonia↗

Role of ammonia in the induction of renal hypertrophy.

Recent experiments from our laboratory have documented the importance of ammonia as a modulator of renal cell growth in vitro. Ammonia induces renal hypertrophy by increasing the rate of protein synthesis and decreasing the rate of protein degradation. These results have led to the hypothesis that an increase in renal ammoniagenesis contributes to renal growth in several seemingly unrelated clinical disorders. In chronic hypokalemia and metabolic acidosis, mitochondrial ammoniagenesis is stimulated directly. During protein loading, uninephrectomy, and diabetes mellitus, renal ammoniagenesis may be stimulated by an increase in single-nephron glomerular filtration rate (SNGFR).

Ammonia↗

pH-induced microtubule-dependent redistribution of late endosomes in neuronal and epithelial cells.

The interaction between late endocytic structures and microtubules in polarized cells was studied using a procedure previously shown to cause microtubule-dependent redistribution of lysosomes in fibroblasts and macrophages (Heuser, J. 1989. J. Cell Biol. 108:855-864). In cultured rat hippocampal neurons, low cytoplasmic pH caused cation-independent mannose-6-phosphate receptor-enriched structures to move out of the cell body and into the processes. In filter grown MDCK cells lowering the cytosolic pH to approximately 6.5 caused late endosomes to move to the base of the cell and this process was shown to be microtubule dependent. Alkalinization caused a shift in distribution towards the apical pole of the cell. The results are consistent with low pH causing the redistribution of late endosomes towards the plus ends of the microtubules. In MDCK cells the microtubules orientated vertically in the cell may play a role in this process. The shape changes that accompanied the redistribution of the late endosomes in MDCK cells were examined by electron microscopy. On low pH treatment fragmentation of the late endosomes was observed whereas after microtubule depolymerization individual late endosomal structures appeared to fuse together. The late endosomes of the MDCK cell appear to be highly pleomorphic and dependent on microtubules for their form and distribution in the cell.

Animals↗

Renal adaptation to dietary phosphate deprivation: role of proximal tubule brush-border membrane fluidity.

With dietary phosphate (Pi) restriction, fluidity of renal proximal tubule brush-border membranes (BBM) and Na-dependent Pi transport (Na-Pi) are increased, suggesting that changes in BBM fluidity are critical for adaptation to Pi restriction. To test this hypothesis, the temporal relationship between Na-Pi transport and changes in BBM fluidity was assessed after Pi deprivation in rats. Renal cortex was obtained from rats fed either a 0.03% (-P) or a 0.6% (+P) Pi diet for 4 h or 7 days, and BBM were prepared. Na-Pi uptake by BBM was measured by use of rapid filtration, and BBM fluidity was assessed by use of the fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH). After 4 h on the diets, Na-Pi uptake was 439 +/- 142 (SD) and 984 +/- 184 pmol.mg protein-1.5 s-1 in +P and -P, respectively (P less than 0.01, n, = 8). Na-dependent proline uptake was unchanged. DPH anisotropy and total cholesterol were similar between groups: 0.204 +/- 0.025 and 0.401 +/- 0.047 nmol/mg protein, respectively, in +P and 0.205 +/- 0.015 and 0.392 +/- 0.037 in -P (P greater than 0.05, n = 8-10). After 7 days, Na-Pi uptake was 841 +/- 291 in +P and 2,168 +/- 848 pmol.mg protein-1.5 s-1 in -P, P less than 0.01, n = 8. DPH anisotropy and BBM cholesterol were 0.175 +/- 0.019 and 443 +/- 132 nmol/mg protein, respectively, in +P and 0.162 +/- 0.020 (n = 8) and 341 +/- 128 (n = 3) in -P (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Adaptation, Physiological↗

H+/base transport in principal cells characterized by confocal fluorescence imaging.

A dual-excitation inverted confocal laser-scanning microscope has been developed for measuring intracellular pH (pHi) using 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF) in individual cells in the isolated perfused cortical collecting tubule (CCT). This new microscope has superior depth discrimination, which eliminates the contribution of fluorescence information from cells outside the plane of focus. pHi was monitored in real time from a spot 0.55 microns in diameter within a single cell. Experiments were performed to examine the apical and basolateral membrane H+/base transport properties of single principal cells. The results indicate that principal cells possess a basolateral membrane Na(+)-independent Cl-/base exchanger, a Na(+)-H+ antiporter, and a Na+/base cotransporter. No evidence was found for an apical membrane Na(+)-independent Cl-/base exchanger. The data provide evidence for base efflux pathways in the principal cell and are compatible with the hypothesis that principal cells contribute importantly to H+/base transport in the CCT. The new methodology described in this report can be applied to other epithelia that are optically heterogeneous in the depth dimension.

Animals↗

Synthesis and characterization of a new fluorescent probe for measuring potassium.

A new fluorescent probe, 6,7-(4-methyl)coumaro-[2.2.2] cryptand, has been developed for measuring K+. This compound was made by fusing [2.2.2] cryptand with 4-methylcoumarin through 2 ethoxy bridges at the 6 and 7 positions. The probe has a fluorescent excitation peak at 340 nm and an emission peak at 420 nm. In aqueous solutions, increasing the K+ concentration from 0 to 10 mM causes the fluorescence intensity to increase by 143%. The dissociation constant (Kd) for K+ in aqueous solutions is 1.9 mM. In 100% methanol, the Kd for K+ decreases to 0.012 mM, making it possible to measure K+ concentrations in the micromolar range. Na+, tetramethylammonium, NH4+, Ca2+, and Mg2+ have a minimal affect on the fluorescence of the probe in the absence of K+. The coefficient of variation for the measurement of K+ by use of this new dye is less than 1%. In this report, the synthetic procedure is described and the spectral properties of the probe are characterized. Experiments are described demonstrating the use of this probe 1) in measuring K+ in aqueous solutions from 0 to 10 mM and in a microfluorometric assay to measure K+ from 0.0005 to 0.003 mM and 2) in monitoring K+ transport in rabbit proximal tubule brush-border membrane vesicles.

Animals↗

Renal ammonia and bicarbonate production in chronic renal failure.

A characteristic feature of chronic renal failure (CRF) is decreased urinary NH4+ excretion and an alteration in systemic acid-base balance. In normal humans, glutamine is an important precursor of urinary NH4+; however, in CRF, renal glutamine extraction is significantly decreased. This finding suggests that alternate sources for urinary NH4+ exist. In addition to being an important precursor of urinary NH4+, the metabolism of glutamine generates alpha-ketoglutarate, an important substrate for new renal HCO3- generation in the proximal tubule. A useful model of the metabolic derangements in human CRF is the subtotally nephrectomized rat. In this review, the ammoniagenic pathways in this model of CRF are characterized with an emphasis on the role of glutathione and gamma-glutamyltransferase. In addition, recent data are presented on the production and partitioning of NH4+ and HCO3- into blood and urine in normal rats and rats with CRF.

Amino Acid Metabolism, Inborn Errors↗

The importance of renal ammonia metabolism to whole body acid-base balance: a reanalysis of the pathophysiology of renal tubular acidosis.

Traditionally, the renal collecting duct has been assigned the dual role of (1) secreting protons derived from dietary metabolism to form luminal NH4+ and titratable acid and (2) generating new HCO3-. This view has recently been challenged. According to current concepts, whole body proton balance is maintained predominantly by the lungs which excrete protons derived from dietary metabolism as the acid anhydride CO2. In the process of excreting CO2, HCO3- is also lost from the body. It is the function of the kidney to generate new HCO3- to replenish this loss. The major site of new HCO3- generation is the proximal tubule rather than the collecting duct. New HCO3- is generated predominantly via the metabolism of organic anions, i.e. alpha-ketoglutarate, citrate, lactate, fatty acids. In the process of generating alpha-ketoglutarate from glutamine, NH4+ is formed. Under normal acid-base conditions, 50% of the NH4+ produced is excreted in the urine, and the remaining 50% is delivered to the renal veins. NH4+ delivered to the renal veins consumes HCO3- during ureagenesis. In the discussion which follows, these new concepts are reviewed and applied to an analysis of the pathophysiology of renal tubular acidosis.

Acid-Base Equilibrium↗

Cl-/base exchange in rat mesangial cells: regulation of intracellular pH.

The present study was designed to determine whether rat glomerular mesangial cells possess Cl- -dependent intracellular pH (pHi) regulatory processes. Rat glomerular mesangial cells were grown to confluence on glass coverslips. Intracellular pH (pHi) was measured with BCECF. Steady state pHi in HCO3- containing solutions was 7.08 +/- 0.03 (N = 13). When extracellular Cl- was acutely removed, pHi increased at a rate of 0.57 +/- 0.03 pH/min units (N = 8), P less than 0.001. DIDS (0.5 mM) significantly decreased the rate of increase in pHi to 0.34 +/- 0.04 pH/min, P less than 0.01. Na+ removal and amiloride (1 mM) did not alter the increase in pHi induced by Cl- removal. Steady state pHi in the absence of Cl- was significantly increased above control, 7.39 +/- 0.02 (N = 7), P less than 0.001. Following the acute alkalinization of pHi by CO2 removal, pHi recovered at a rate of 0.07 +/- 0.01 pH/min (N = 9). In the absence of Cl-, the pHi recovery rate was significantly decreased to 0.01 +/- 0.008 pH/min (N = 5), P less than 0.01. DIDS (0.5 mM) significantly decreased the rate of pHi recovery to 0.02 +/- 0.01 pH/min (N = 5), P less than 0.01. Na+ removal and amiloride (1 mM) had no effect on the rate of pHi recovery following acute alkaline loading.(ABSTRACT TRUNCATED AT 250 WORDS)

Acid-Base Equilibrium↗

Basolateral membrane Na+/H+ antiport, Na+/base cotransport, and Na+-independent Cl-/base exchange in the rabbit S3 proximal tubule.

The basolateral membrane Na+ and Cl(-)-dependent acid-base transport processes were studied in the isolated perfused rabbit S3 proximal straight tubule. Intracellular pH (pHi) was measured with 2'7'-biscarboxyethyl-5,6-carboxyfluorescein (BCECF) and a microfluorometer coupled to the tubule perfusion apparatus. Reduction of basolateral HCO3- from 25 to 5 mM caused pHi to decrease at a rate of 0.81 pH/min. Approximately 50% of this rate was Na+-dependent, 30% Cl(-)-dependent and 20% Na+ and Cl(-)-independent. Two basolateral Na+-dependent acid base transport pathways were detected: (a) an amiloride-sensitive Na+/H+ antiporter and (b) a stilbene-sensitive Na+/base cotransporter. No evidence was found for a Na+-dependent Cl-/base exchanger. The Cl(-)-dependent component of basolateral base efflux was mediated by a stilbene-sensitive Na+-independent Cl-/base exchange pathway. The results suggest that the acid base transport pathways of the basolateral membrane of the S3 proximal tubule differ from more proximal nephron segments.

4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfo↗

Induction of hypertrophy in cultured proximal tubule cells by extracellular NH4Cl.

Ammonia production increases in several models of renal hypertrophy in vivo. The present study was designed to determine whether ammonia can directly modulate the growth of renal cells in the absence of a change in extracellular acidity. In serum-free media NH4Cl (0-20 mM) caused JTC cells and a primary culture of rabbit proximal tubule cells to hypertrophy (increase in cell protein content) in a dose-dependent fashion without a change in DNA synthesis. Studies in JTC cells revealed that the cell protein content increased as a result of both an increase in protein synthesis and a decrease in protein degradation. Total cell RNA content and ribosome number increased after NH4Cl exposure and the cell content of the lysosomal enzymes cathepsin B and L decreased. Inhibition of the Na+/H+ antiporter with amiloride did not prevent the hypertrophic response induced by NH4Cl. The results indicate that ammonia is an important modulator of renal cell growth and that hypertrophy can occur in the absence of functioning Na+/H+ antiport activity.

Amiloride↗

Apical and basolateral Na+/H+ exchange in the rabbit outer medullary thin descending limb of Henle: role in intracellular pH regulation.

The present study was designed to investigate the apical and basolateral transport processes responsible for intracellular pH regulation in the thin descending limb of Henle. Rabbit thin descending limbs of long-loop nephrons were perfused in vitro and intracellular pH (pHi) was measured using BCECF. Steady-state pHi in HEPES buffered solutions (pH 7.4) was 7.18 +/- 0.03. Following the removal of luminal Na+, pHi decreased at a rate of 1.96 +/- 0.37 pH/min. In the presence of luminal amiloride (1 mM), the rate of decrease of pHi was significantly less, 0.73 +/- 0.18 pH/min. Steady-state pHi decreased 0.18 pH units following the addition of amiloride (1 mM) to the lumen (Na+ 140 mM lumen and bath). When Na+ was removed from the basolateral side of the tubule, pHi decreased at a rate of 0.49 +/- 0.05 pH/min. The rate of decrease of pHi was significantly less in the presence of 1 mM basolateral amiloride, 0.29 +/- 0.04 pH/min. Addition of 1 mM amiloride to the basolateral side (Na+ 140 mM lumen and bath) caused steady-state pHi to decrease significantly by 0.06 pH units. When pHi was acutely decreased to 5.87 +/- 0.02 following NH4Cl removal (lumen, bath), pHi failed to recover in the absence of Na+ (lumen, bath). Addition of 140 mM Na+ to the lumen caused pHi to recover at a rate of 2.17 +/- 0.59 pH/min. The rate of pHi recovery was inhibited 93% by 1 mM luminal amiloride. When 140 mM Na+ was added to the basolateral side, pHi recovered only partially at 0.38 +/- 0.07 pH/min. Addition of 1 mM basolateral amiloride inhibited the recovery of pHi by 97%. The results demonstrate that the rabbit thin descending limb of long-loop nephrons possesses apical and basolateral Na+/N+ antiporters. In the steady state, the rate of Na+-dependent H+ flux across the apical antiporter exceeds the rate of Na+-dependent H+ flux via the basolateral antiporter. Recovery of pHi following acute intracellular acidification is Na+ dependent and mediated primarily by the luminal antiporter.

Animals↗

NH3 permeation through the apical membrane of MDCK cells is via a lipid pathway.

The pathway for NH3 permeation across the apical membrane of MDCK cells was determined by measuring the effect of membrane fluidizing agents, protein reactive agents, and temperature on cellular NH3 influx. The rate of NH3 influx was calculated from the time course of increase in intracellular pH (pHi), measured with 2,7-biscarboxyethyl-5(6)-carboxyfluorescein, when MDCK cell monolayers were exposed to NH4Cl. The apical membrane NH3 permeability was 7.13 +/- 0.37 x 10(-3) cm/s (n = 12) at 37 degrees C and 1.23 +/- 0.07 x 10(-3) cm/s (n = 7) at 18 degrees C. In comparison, apical membrane permeability at 37 degrees C to the weak acids, valeric acid and acetic acid, were 1.39 +/- 0.11 x 10(-2) cm/s (n = 4) and 6.93 +/- 0.11 x 10(-3) cm/s (n = 4), respectively. The activation energy for NH3 permeation was 15.0 +/- 1.0 kcal/mol (17.5 degrees C-37.5 degrees C). In the presence of the membrane fluidizing agents, heptanol or chloroform, NH3 permeability increased in a dose-dependent manner. Heptanol (15 mM) significantly decreased the activation energy for NH3 permeation to 4.4 +/- 0.6 kcal/mol, P less than 0.001. The carboxyl reactive agent (1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide metho-p-toluensulfonic acid 1 mM), aminoreactive agents (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid 50 microM; picrylsulphonic acid 1 mM), the sulphydryl reactive agent (p-chloromercuriphenylsulfonic acid 1 mM), and the nonspecific membrane protein cleaving agent pronase (1 mg/ml) had no effect on the NH3 influx. The results suggest that NH3 permeates the plasma membrane of MDCK cells via a lipid pathway.

4-Chloromercuribenzenesulfonate↗

Na+-independent Cl(-)-HCO-3- exchange in Madin-Darby canine kidney cells. Role in intracellular pH regulation.

The role of plasma membrane Cl(-)-HCO-3-exchange in regulating intracellular pH (pHi) was examined in Madin-Darby canine kidney cell monolayers. In cells bathed in 25 mM HCO-3, pH 7.4, steady state pHi was 7.10 +/- 0.03 (n = 14) measured with the fluorescent pH probe 2',7'-biscarboxyethyl-5,6-carboxyfluorescein. Following acute alkaline loading, pHi recovered exponentially in approximately 4 min. The recovery rate was significantly decreased by Cl- or HCO-3 removal and in the presence of 50 microM 4,4'-diisothiocyano-2,2'-disulfonic stilbene (DIDS). Na+ removal or 10(-3) M amiloride did not inhibit the pHi recovery rate after an acute alkaline load. Following acute intracellular acidification, the pHi recovery rate was significantly inhibited by 10(-3) M amiloride but was not altered by Cl- removal or 50 microM DIDS. At an extracellular pH (pHo) of 7.4, pHi remained unchanged when the cells were bathed in either Cl- free media, HCO-3 free media, or in the presence of 50 microM DIDS. As pHo was increased to 8.0, steady state pHi was significantly greater than control in Cl(-)-free media and in the presence of 50 microM DIDS. It is concluded that Madin-Darby canine kidney cells possess a Na+-independent Cl(-)-HCO-3 exchanger with a Km for external Cl- of approximately 6 mM. The exchanger plays an important role in pHi regulation following an elevation of pHi above approximately 7.1. Recovery of pHi following intracellular acidification is mediated by the Na+/H+ antiporter and not the anion exchanger.

Animals↗

Measurement of intracellular Ca2+ activity in Necturus gallbladder.

To clarify the effects of Ca2+-free solutions on the electrophysiological properties of leaky epithelia, Necturus gallbladder was mounted in an Ussing-type chamber and its mucosal surface exposed to Ca2+-free EGTA (2 mM) Ringer. Lateral-space width was controlled by a -3-cmH2O pressure gradient on the serosal outflow. Transepithelial potential difference and resistance were monitored while cell membrane potential and intracellular Ca2+ activity (Ca2+i) were determined with conventional and Ca2+-sensitive microelectrodes. Ca2+i averaged 183 +/- 27 nM (n = 15). Reduction of mucosal Ca2+ activity to approximately 500 nM reversibly lowered transepithelial resistance while cell membrane potential remained unaltered and fractional membrane resistance increased from 0.77 +/- 0.01 to 0.83 +/- 0.02 (P less than 0.01, n = 5). In five gallbladders mucosal Ca2+ reduction induced a significant drop in Ca2+i from 133 +/- 26 to 77 +/- 20 nM (P less than 0.01, n = 5) while transepithelial resistance fell from 125 +/- 27 to 107 +/- 24 omega X cm2 (P less than 0.01). These results indicate that transepithelial resistance decrements observed during exposure to Ca2+-free solutions stem from a reversible increase in tight-junctional but not cell membrane permeability and that this effect is associated with a fall in intracellular Ca2+ activity.

Animals↗