bluuebunny/crossref_metadata_2025 huggingface dataset:
Sodium-phosphate cotransport in OK cells: inhibition by PTH and "adaptation" to low phosphate
Authors: Quamme G. , Biber J. , Murer H.
Published: December 2017

Sodium-phosphate (Na-Pi) cotransport is principally regulated by parathyroid hormone (PTH) and by the intrinsic ability to “adapt” to ambient phosphate concentration. In the present study, these two control mechanisms were examined in a cloned opossum kidney (OK) cell line. PTH inhibited Na-Pi cotransport, half-maximal inhibition at 5 x 10(-12) M, by fractionally similar amounts irrespective of the initial transport rates predetermined by “adaptation” to media phosphate concentration. At maximal concentrations of PTH (10(-8) M), the residual Na-Pi cotransport activity was higher in cells exposed to low-phosphate media. Cells preexposed to PTH (10(-8) M) or dibutyryl adenosine 3'-5'-cyclic monophosphate (DBcAMP) (10(-5) M) and forskolin (10(-5) M) increase transport (adaptation) by fractionally similar amounts as control cells for any given external phosphate concentration. The protein kinase C inhibitor, staurosporine, prevented PTH action but did not alter the ability to adapt Na-Pi cotransport in response to low media phosphate concentration. These data support the notion that regulation of Na-Pi cotransport by PTH and the adaptive response to available media phosphate concentration are distinct regulatory control mechanisms.
Effect of protein intake on the autoregulation of renal blood flow
Authors: Murray B. M. , Brown G. P.
Published: December 2017

Autoregulatory ability was evaluated in male Sprague-Dawley rats (200-250 g) fed either low protein (LP, 6%), high protein (HP, 50%), or standard rat chow (CON) for 2 wk. Renal perfusion pressure (RPP) was first raised above normal by bilateral ligation of the carotid arteries. LP rats exhibited normal autoregulatory behavior between 130 and 100 mmHg, whereas autoregulation in HP rats was impaired (autoregulation factor 100-130; HP 0.86 +/- 0.16, LP 0.38 +/- 0.07, and CON 0.39 +/- 0.04). Autoregulation studies were repeated in HP- and LP-fed rats after administration of the cyclooxygenase inhibitors, meclofenamate and piroxicam. Cyclooxygenase inhibition did not significantly affect base-line hemodynamics but did restore the ability of the HP rat to autoregulate renal blood flow (RBF) at high RPP. (Autoregulation factor between 100 and 130 mmHg; HP 0.29 +/- 0.10, LP 0.19 +/- 0.07, P, NS). Thus an HP diet resulted in impaired autoregulation of RBF at high RPP, an effect that appears to be mediated by excessive production of vasodilatory prostaglandins.
Cytosolic free calcium regulation in renal tubules from spontaneously hypertensive rats
Authors: Jacobs W. R. , Ferrari C. M. , Brazy P. C. , Mandel L. J.
Published: December 2017

Cytosolic free calcium (Caf) was measured in isolated renal cortical tubules from spontaneously hypertensive rats (SHR) and age-matched control Wistar-Kyoto (WKY) rats to evaluate whether the onset of hypertension is associated with a change in Caf. At the same time, the cellular mechanism by which differences in Caf occur between these two strains of rats was examined. Caf was significantly lower in renal tubules from 4- to 5-wk (142 +/- 6 vs. 187 +/- 15 nM), 6- to 7-wk (138 +/- 15 vs. 187 +/- 8 nM), and 8- to 9-wk-old (161 +/- 5 vs. 216 +/- 9 nM) SHR compared with age-matched WKY. The lower Caf in SHR tubules was considered to be the result of either an increase in Ca efflux or a reduction in Ca permeability. To the extent that metabolic inhibitors increased Caf but did not alter the difference between SHR and WKY, a primary difference in Caf efflux was excluded. Conversely, when Ca permeability was altered, either with Ca ionophores or incubation in Ca-free medium, Caf changed in the appropriate direction and the difference between SHR and WKY was no longer apparent. These results demonstrate that 1) the previously reported increase in Caf in circulating cells is not a universal feature of hypertension and 2) the lower Caf in renal tubules from SHR appears to be related to a lower Ca permeability. Whether the differences in Caf between SHR and WKY is a permissive factor for the renal contribution to hypertension remains to be elucidated.
Aromatic L-amino acid decarboxylase activity along the rat nephron
Authors: Hayashi M. , Yamaji Y. , Kitajima W. , Saruta T.
Published: December 2017

Extraneural dopamine is thought to be synthesized by an aromatic L-amino acid decarboxylase (L-AADC) activity in tubular cells. However, the previous histochemical studies of this enzyme's localization in the nephron were not consistent. To determine the localization of L-AADC and whether changes in Na intake regulate this enzyme, L-AADC was measured in microdissected nephron segments from rat kidneys. Dopamine formed by isolated tubules incubated with exogenous L-dopa was quantitated by high-performance liquid chromatography (HPLC) and with the more sensitive radioenzyme assay (REA). L-AADC activity was present only in proximal convoluted (PCT, 208 +/- 19 ng.cm-1.h-1) and proximal straight tubules (PST, 81 +/- 9 ng.cm-1.h-1), whereas no significant activity was detected in other nephron segments by either HPLC or REA. Maximal velocity (Vmax) of L-AADC in a low-salt diet group (246 +/- 4 ng.cm-1.h-1) showed a small but a significant decrease (P less than 0.05) compared with control and high-salt diet groups (311 +/- 6 and 293 +/- 4 ng.cm-1.h-1, respectively), whereas the apparent Michaelis constant (Km) was similar in these three groups. These results show that L-AADC is present only in the PCT and PST of the rat nephron, and suggest that the changes in L-AADC activity may contribute to the regulation of extraneural dopamine production in the kidney during low-salt intake.
Distal tubular acidification in the remnant kidney
Authors: Kunau R. T. , Walker K. A.
Published: December 2017

The present studies examined the effect of three-fourths nephrectomy on the rate of acidification, i.e., total CO2 (tCO2) absorption (JtCO2) in the superficial distal tubule of the rat. Total glomerular filtration rate following three-fourths nephrectomy was 1.29 +/- 0.06 vs. 3.29 +/- 0.08 ml/min in sham controls, P less than 0.001. Systemic acid-base parameters were the same in both groups, but urine pH was lower in nephrectomized rats. In vivo microperfusion with identical isohydric solutions revealed that the JtCO2, fluid absorption (Jv), lumen-negative transepithelial potential difference (VT) were all significantly greater in the distal tubule of remnant kidneys. As the relative increase in Jv exceeded JtCO2, the perfusate tCO2 concentration increased markedly in remnant kidney distal tubules from 30.3 +/- 0.59 to 39.9 +/- 1.73 mM. To determine if the increase in tCO2 concentration accounted for the difference in JtCO2, a second control group was studied using a perfusate tCO2 concentration of 39.6 +/- 0.79 mM. Distal tubular JtCO2, Jv, and VT were significantly less in this control group than in the remnant kidney group. In separate studies, 10(-4) M amiloride was added to the perfusate used in remnant kidneys and controls studied with the elevated perfusate tCO2 concentration. The addition of 10(-4) M amiloride to the perfusate reduced VT and JtCO2. At identical values for VT, JtCO2 was higher in the distal tubule of remnant kidneys than in controls. We conclude the following. 1) The rate of acidification is increased in the distal tubule of remnant kidneys.(ABSTRACT TRUNCATED AT 250 WORDS)
Stimulation by thyroid hormone of Na+-H+ exchange activity in cultured opossum kidney cells
Authors: Yonemura K. , Cheng L. , Sacktor B. , Kinsella J. L.
Published: December 2017

Cultured opossum kidney (OK) cells were used to determine whether thyroid hormone has a direct stimulatory effect on Na(+)-H+ exchange activity in an intact cellular preparation. Na+ uptake or intracellular pH recovery was measured in confluent monolayer cells following acid loading with NH4Cl. Triiodo-L-thyronine (T3) had no effect on cell number or protein and DNA contents but stimulated amiloride-sensitive Na+ uptake in a dose- and time-dependent manner. Maximal stimulation of Na+ uptake was observed at 10(-7) M T3 and 10(-6) M L-thyroxine (T4) with half-maximal effects at 10(-9) M T3 and 3 x 10(-8) M T4. The T3 specific binding capacity of OK cells was 96 +/- 15 fmol/mg DNA with a KD of 1.1 +/- 0.2 x 10(-9) M T3. Neither T3 nor T4 had any effect on amiloride-insensitive Na+ uptake. In kinetic studies of Na+ uptake, T3 increased the Vmax from 123 +/- 22 to 157 +/- 24 nmol.mg-1.min-1 without changing the Michaelis-Menten kinetics (Km) for Na+ (21 +/- 1 in control and 22 +/- 4 mM in T3-treated cells). Studies of intracellular pH (pHi) showed that the resting pHi and the buffering capacity were unaffected by T3. However, after an acid load, OK cells treated with T3 exhibited a greater rate of Na(+)-dependent pH recovery than untreated control cells. These results indicate that thyroid hormone can stimulate Na(+)-H+ exchange activity directly in renal cell line without apparent changes in pHi, cellular hypertrophy, or hyperplasia.
Intracellular pH regulation in rabbit S3 proximal tubule: basolateral Cl-HCO3 exchange and Na-HCO3 cotransport
Authors: Nakhoul N. L. , Chen L. K. , Boron W. F.
Published: December 2017

We studied the role of basolateral HCO3- transport in the regulation of intracellular pH (pHi) in the isolated perfused S3 segment of the rabbit proximal tubule. pHi was calculated from absorbance spectra of the pH-sensitive dye dimethylcarboxyfluorescein. Solutions were normally buffered to pH 7.4 at 37 degrees C with 25 mM HCO3- 5% CO2. pHi fell by approximately 0.17 when luminal [HCO3-] was lowered to 5 mM at fixed PCO2 (i.e., reducing pH to 6.8) but by approximately 0.42 when [HCO3-] in the bath (i.e., basolateral solution) was lowered to 5 mM. The pHi decrease elicited by reducing bath [HCO3-] was substantially reduced by removal of Cl- or Na+, suggesting that components of basolateral HCO3- transport are Cl- and/or Na+ dependent. We tested for the presence of basolateral Cl-HCO3 exchange by removing bath Cl-. This caused pHi to increase by approximately 0.23, with an initial rate of approximately 100 X 10(-4) pH/s. Although the initial rate of this pHi increase was not reduced by removing Na+ bilaterally, it was substantially lowered by the nominal removal of HCO3- from bath and lumen or by the addition of 0.1 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) to the bath. The results thus suggest that a Na-independent Cl-HCO3 exchanger is present at the basolateral membrane. We tested for the presence of basolateral Na-HCO3 cotransport by removing bath Na+. This caused pHi to fall reversibly by approximately 0.26 with initial rates of pHi decline and recovery being approximately 30 and approximately 41 X 10(-4) pH/s, respectively. Although the bilateral removal of Cl- had no effect on these rates, the nominal removal of HCO3- or the presence of DIDS substantially slowed the pHi changes. Thus, in addition to a Cl-HCO3 exchanger, the basolateral membrane of the S3 proximal tubule also appears to possess a Na-HCO3 cotransport mechanism. The data do not rule out the possibility of other basolateral HCO3- transporters.
Glomerulotubular balance in a mathematical model of the proximal nephron
Authors: Weinstein A. M.
Published: December 2017

A nonelectrolyte model of proximal tubule epithelium has been extended by the inclusion of a compliant tight junction. Here "compliance" signifies that both the junctional salt and water permeability increase and the salt reflection coefficient decreases in response to small pressure differences from lateral interspace to tubule lumen. In previous models of rat proximal tubule, there has been virtually no sensitivity of isotonic salt transport to changes in peritubular oncotic force. With the inclusion of junctional compliance, decreases in peritubular protein can open the junction and produce a secretory salt flux. Thus the model can represent the "backflux hypothesis," as it was originally put forth (J. E. Lewy and E. E. Windhager, Am. J. Physiol. 214: 943-954, 1968). Additional calculations, simulating a tight junction with negligible water permeability, reveal that the quantitative impact of peritubular protein can be realized whether or not there is substantial junctional water flux. The epithelial model of proximal tubule has also been incorporated into a model of the proximal nephron, complete with glomerulus, peritubular capillary, and interstitium. The interstitial compartment is well mixed and interstitial pressure and osmolality are determined iteratively to achieve balance between tubule reabsorption and capillary uptake. For this model, two domains of operation are identified. When interstitial pressures are low, junctions are closed, and filtration fraction has no effect on proximal reabsorption. When interstitial pressures are relatively elevated, epithelial junctions are open, and proximal salt reabsorption changes in proportion to changes in filtration fraction. In neither domain, however, does the model tubule augment salt flux with isolated increases in luminal flow rate (at constant filtration fraction). The absence of a separate effect of tubule fluid flow on salt transport precludes perfect glomerulotubular balance.
Renal actions of atrial natriuretic factor: modulation of effect by changes in sodium status and aldosterone
Authors: Yates N. A. , Coghlan J. P. , Murphy G. J. , Scoggins B. A. , McDougall J. G.
Published: December 2017

To examine the effect of changes in Na status on the renal response to atrial natriuretic factor (ANF), human ANF-(99-126) was infused into the renal artery of conscious sheep while Na replete, Na depleted, or Na loaded. The natriuretic response to ANF was attenuated during Na depletion and enhanced in Na-loaded animals. To demonstrate that the enhancement or attenuation of response was related to Na status rather than to the initial level of Na excretion, aldosterone was infused into the renal artery for 2 h to decrease Na excretion to a level inappropriately low for the animal's Na status (and not different from Na-depleted animals), and they were again challenged with ANF. Their response to ANF, however, was not significantly different from that in normal Na-replete animals but was significantly greater than that observed in Na-depleted animals. Similarly, Na-loaded animals treated with aldosterone had control Na excretion in the Na-replete range; however, their response to ANF was not significantly different from that of Na-loaded animals. The response to ANF was enhanced in sheep treated with aldosterone for 48 h, consistent with Na retention and hypervolemia, secondary to aldosterone treatment. The study demonstrates that Na status or some associated physiological parameter is an important determinant of the natriuretic response to ANF. The present series of experiments demonstrate that changes in aldosterone levels per se are not a determinant of the natriuretic response to ANF.
A micropuncture study of proximal tubular transport of lithium during osmotic diuresis
Authors: Leyssac P. P. , Holstein-Rathlou N. H. , Skott P. , Alfrey A. C.
Published: December 2017

Lithium and sodium are normally reabsorbed in parallel with water by the renal proximal tubule whereby their tubular fluid-to-plasma concentration ratios (TF/P) remain close to unity throughout the proximal convoluted segment. During osmotic diuresis, the late proximal (TF/P)Na is known to decrease. The present experiments were undertaken to study whether the late proximal TF/P for Li decreases like that of Na during osmotic diuresis. Data were obtained in a control period (C) and in two successive periods during mannitol diuresis (P1, P2). Glomerular filtration rate decreased gradually during osmotic diuresis [1,147 +/- 32 (C) to 950 +/- 22 (P2) microliters.min-1.g kidney wt (KW)-1, P less than 0.01), whereas Li clearance (CLi) increased [377 +/- 21 (C) to 469 +/- 28 (P1, P less than 0.01) and 408 +/- 25 (P2, NS) microliters.min-1.gKW-1, respectively]. (TF/P)Na decreased from 0.98 +/- 0.02 (C) to 0.89 +/- 0.02 (P less than 0.01) and 0.90 +/- 0.04 (P less than 0.05) (P1 and P2, respectively). (TF/P)Li was close to, but significantly higher than, unity during control (1.09 +/- 0.02), but, in contrast to (TF/P)Na, it did not decrease significantly during osmotic diuresis. Delivery of water from the proximal straight segment, as estimated from CLi/CIn, increased in proportion to the delivery from the convoluted segment, as estimated from 1/(TF/P)In. It is concluded that the proximal net Li reabsorption follows more closely the net water reabsorption than the net Na reabsorption during osmotic diuresis.
Cytochrome P-450 arachidonate metabolites in rat kidney: characterization and hemodynamic responses
Authors: Takahashi K. , Capdevila J. , Karara A. , Falck J. R. , Jacobson H. R. , Badr K. F.
Published: December 2017

Rat kidney cortical and medullary microsomal fractions catalyzed cytochrome P-450-linked metabolism of arachidonic acid (AA) to epoxyeicosatrienoic acids (EETs) (56 +/- 6% of total products in cortex and 10% in medulla) and 19- and 20-hydroxyeicosatetraenoic acids (19- and 20-OHAA) (36 +/- 4% in cortex and 90% in medulla). In addition, endogenous renal generation of EETs was established by negative ion-chemical ionization mass spectrometry. The total amount of EETs present in the rat kidney was approximately 1 microgram/g wet tissue. The responses to renal arterial administration of 20-OHAA and 5,6-EET were evaluated in anesthetized euvolemic rats. 20-OHAA resulted in ipsilateral dose-dependent natriuresis without affecting systemic or renal hemodynamics or glomerular filtration rate (GFR). Equimolar doses of 5,6-EET resulted in dose-dependent renal vasoconstriction and reduced GFR but were without effect on arterial pressure or sodium excretion. During cyclooxygenase inhibition, 5,6-EET caused renal vasodilatation and augmentation of GFR. These data establish the capacity of rat kidney to metabolize AA through cytochrome P-450-dependent oxygenases and demonstrate the endogenous formation of the resulting eicosanoids. Monooxygenase and epoxygenase products exert effects on renal blood flow, GFR, and urinary sodium excretion rate, suggesting their potential relevance in the regulation of renal function.
Attenuation of enhanced tubuloglomerular feedback activity in SHR by renal denervation
Authors: Takabatake T. , Ushiogi Y. , Ohta K. , Hattori N.
Published: December 2017

We evaluated the effect of acute unilateral renal denervation (DNX) on the tubuloglomerular feedback (TGF) mechanism in Inactin-anesthetized hydropenic male 8- to 10-wk-old spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). SHR had higher mean arterial pressure (MAP, 28%) and renal vascular resistance (RVR, 35%), whereas renal blood flow (RBF), glomerular filtration rate (GFR), urine flow, and sodium excretion were similar. DNX in SHR did not change MAP but decreased RVR (26%) and increased RBF (29%), GFR (16%), urine flow (52%), and sodium excretion (431%). DNX did not affect these in WKY. Loop of Henle perfusion with Ringer solution reduced early proximal flow rate (EPFR) in SHR more than in WKY; significantly different at a loop flow of 20 nl/min (9.8 +/- 0.7 vs. 6.5 +/- 0.7 nl/min). DNX in SHR increased the nonperfused EPFR from 25.6 +/- 1.1 to 31.7 +/- 1.3 nl/min and reduced TGF responses during perfusion at both 20 nl/min (9.8 +/- 0.7 vs. 4.4 +/- 0.7 nl/min) and 40 nl/min (14.2 +/- 1.1 vs. 10.4 +/- 0.7 nl/min). TGF sensitivity was attenuated by DNX, as indicated by reduced maximum reactivity (-0.89 +/- 0.14 to -0.36 +/- 0.07) and increased turning point (16.5 +/- 0.9 to 25.2 +/- 2.9 nl/min). TGF responses in WKY were not influenced by DNX. Sham denervation did not alter renal hemodynamics and TGF. These results indicate that renal nerves exert a tonic influence on the renal vasculature and the TGF system in SHR but not in WKY. Enhanced TGF responsiveness may be involved in volume retention and in the maintenance of hypertension in SHR.
Renal vascular response to amino acids: effect of pancreatectomy
Authors: Premen A. J. , Powell D. A. , Carroll R. G. , Dobbins D. E.
Published: December 2017

We ascertained the importance of glucagon in modulating the renal hemodynamic response to amino acid (AA) infusion in anesthetized dogs. In controls (n = 6), AAs (L-serine, alanine, and proline; 0.051 mmol.kg-1.min-1 iv) elevated renal blood flow (RBF) and glomerular filtration rate (GFR) by 35 and 34%, respectively, while elevating arterial plasma glucagon-like immunoreactivity (AGLI) by 96 pmol/l. In control pancreatectomized (PX) dogs (n = 6), all parameters remained at control values over 2 h. In PX dogs, AAs (n = 6) failed to reproduce the renal hemodynamic and AGLI responses elicited by AAs in controls. In PX dogs infused with AAs, replacement of AGLI (n = 6) to an incremental plasma level of 111 pmol/l, a level no different than that produced by AAs in controls, elevated RBF and GFR by 25 and 26.5%, respectively. These hemodynamic responses were 71 and 78%, respectively, of the total responses elicited by AAs in controls. In PX dogs infused with glucagon alone (0.86 pmol.kg-1.min-1; n = 6), an incremental change in AGLI of 112 pmol/l was accompanied by only small increases in RBF and GFR (9%). These data suggest the importance of glucagon in modulating the renal hyperemia and hyperfiltration ascribed to AA infusion in anesthetized dogs.
Na(+)-H+ exchange, but not Na(+)-K(+)-ATPase, is present in endosome-enriched microsomes from rabbit renal cortex
Authors: Hilden S. A. , Ghoshroy K. B. , Madias N. E.
Published: December 2017

Previous studies have demonstrated a Na(+)-dependent decrease in the ATP-generated acidification of endosomes and have attributed it to the presence of either a Na(+)-H+ exchanger or a Na(+)-K(+)-adenosinetriphosphatase (ATPase) in parallel with the vacuolar H(+)-ATPase. In the present study we have examined the possibility that both of these two Na+ transporters might be present in endosome-enriched microsomes isolated from rabbit renal cortex. After the establishment of a stable pH gradient by ATP in this preparation, addition of Na+ induced a decrease in the pH gradient. Expression of this effect of Na+ did not require the presence of ATP or K+. Choline and K+ had no effect on the ATP-dependent pH gradient, but addition of Li+ caused a small reduction in the pH gradient. Amiloride, ouabain, and vanadate had no effect on the Na(+)-induced dissipation of the ATP-driven pH gradient. In addition, a pH gradient-dependent 22Na+ uptake by the endosomal vesicles that was insensitive to amiloride, ouabain, or vanadate was demonstrated. These results provide evidence against the presence of a Na(+)-K(+)-ATPase in endosome-enriched microsomes from the renal cortex and support the existence of an amiloride-insensitive Na(+)-H+ exchanger in parallel with the vacuolar H(+)-ATPase. This endosomal Na(+)-H+ exchanger might have important implications for the regulation of vacuolar H(+)-ATPase activity as well as proximal tubule acidification.
Cortical collecting duct Na-K pump in obstructive nephropathy
Authors: Kimura H. , Mujais S. K.
Published: December 2017

The present study examined the alterations in the cortical collecting tubule (CCT) Na-K pump that occur after unilateral ureteral obstruction and their consequences on electrolyte excretion. In male Sprague-Dawley rats, unilateral ureteral ligation led to a progressive decrease in intact CCT Na-K pump in situ turnover worsening with the duration of the obstruction: control, 20.1 +/- 0.4; obstructed kidney: 3 h, 14.6 +/- 0.3; 12 h, 12.7 +/- 0.6; 24 h, 12.8 +/- 0.5; 48 h, 11.6 +/- 0.5; and 96 h, 10.6 +/- 0.4 pmol Rb.mm-1.min-1 (all P less than 0.001 vs. control). CCT diameter increased with the duration of obstruction. Release of ureteral obstruction was associated with restitution of pump turnover rate. With 3 h of obstruction, recovery of pump in situ turnover was complete (19.7 +/- 0.4 pmol Rb.mm-1.min-1) by 24 h after release. With more prolonged obstruction (24 h) recovery was partial by 24 h postrelease (16.2 +/- 0.5 pmol Rb.mm-1.min-1) and complete (19.8 +/- 0.7 pmol Rb.mm-1.min-1) by 48 h, suggesting a delay in recovery without the occurrence of irreversible damage. The impairment in Na-K pump in situ turnover was paralleled by an impairment in the ability of the obstructed kidney to excrete an acute potassium load. This parallelism of functional and biochemical studies favors the notion that impairment of CCT Na-K pump in situ turnover contributes significantly to the abnormal potassium excretion that accompanies obstructive damage.
A dynamic model of the tubuloglomerular feedback mechanism
Authors: Holstein-Rathlou N. H. , Marsh D. J.
Published: December 2017

We have reported oscillations in proximal tubular pressure and flow and in distal tubular pressure and chloride concentration in halothane-anesthetized Sprague-Dawley rats. These variables oscillated at the same frequency in each animal, approximately 35 mHz, but were out of phase with each other. We suggested that the oscillation arises within the tubuloglomerular feedback (TGF) system. As a test of this hypothesis, we have now developed a dynamic model to determine whether it can simulate the measured frequency and phase relationships with a realistic set of parameters. The model includes a detailed representation of pressure and flow in the tubules based on a reduced version of the Navier-Stokes equations. The NaCl concentration at the macula densa was used as the signal to the TGF mechanism. The tubular NaCl concentration was modeled by a partial differential equation based on conservation of mass. For a realistic set of parameter values the model accurately predicted oscillations with the same frequency and phase relationships among the oscillating variables as was found experimentally. Moreover, tubular NaCl handling significantly influenced the dynamic properties of the TGF system. Thus the model predicted a substantial phase shift of the NaCl concentration relative to the flow oscillation at the macula densa. The results are consistent with the hypothesis that the oscillations are caused by the TGF mechanism. The results further support the notion that the delays and damping caused by the tubule are responsible for the limited high-frequency response of renal autoregulation.
Chloride transport across the basolateral membrane of rabbit proximal convoluted tubules
Authors: Ishibashi K. , Rector F. C. , Berry C. A.
Published: December 2017

To examine the basolateral Cl transport mechanisms of proximal convoluted tubules (PCT), intracellular Cl activity (AiCl) was measured with double-barreled Cl-selective microelectrodes. When rabbit PCT were perfused in vitro with high Cl, low HCO3, and bathed with ultrafiltrate-like solutions, AiCl was 29.9 +/- 0.4 mM and basolateral membrane voltage (Vbl) was -47.7 +/- 0.4 mV (n = 247). Possible basolateral Cl transport mechanisms that we examined were as follows: Cl conductance, KCl cotransport, and Na-dependent Cl-HCO3 exchange. Cl conductance was negligible, since the voltage clamp of Vbl to 30 mV above and below the spontaneous Vbl did not change AiCl even in the absence of luminal Cl. KCl cotransport was suggested by 1) increasing bath K, increased AiCl, and 2) decreasing bath K decreased AiCl. KCl cotransport was Na independent and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS), barium, and furosemide insensitive. Na-dependent Cl-HCO3 exchange was suggested by 1) bath HCO3 reduction increased AiCl, which was greatly inhibited by bath Na removal or bath SITS, and 2) bath Na removal increased AiCl, which was completely blocked by bath SITS. We conclude that 1) Cl conductance is negligibly small at the basolateral membrane and 2) SITS-insensitive KCl cotransport and SITS-sensitive Na-dependent Cl-HCO3 exchange are present at the basolateral membrane.
Vitamin D-induced chronic hypercalcemia inhibits thick ascending limb NaCl reabsorption in vivo
Authors: Peterson L. N.
Published: December 2017

Chronic hypercalcemia (HC) induced by dihydrotachysterol (DHT) is associated with a nephrogenic concentrating defect. The purpose of the present study is to assess the effects of HC on thick ascending limb (TAL) NaCl reabsorption in vivo by use of micro stop-flow technique. Feeding DHT (4.25 mg/kg diet) to rats was associated with an increase in plasma [Ca] within 24 h from 2.21 +/- 0.049 to 2.71 +/- 0.047 mM (P less than 0.001), which remained elevated during the 7-day period of study. Ambient plasma arginine vasopressin (AVP) in polydipsic HC rats was 3.10 +/- 0.605 pg/ml, a value not different from that measured in pair-fed control rats (1.82 +/- 0.260 pg/ml). A urine-concentrating defect developed after 3 days and occurred without nephrocalcinosis or reduced glomerular filtration rate (GFR). The estimated TAL [NaCl] after all stop-flow periods in HC rats did not decrease to the same extent as in controls. The rate constant of NaCl reabsorption derived from linear regression analysis of the ln[NaCl] vs. time for 0-15 s was significantly reduced by 25% in the HC nephrons. In addition, the minimum [NaCl] established after 30-60 s of contact time increased by 36% in HC nephrons. These data provide evidence that reduced TAL NaCl reabsorption, and not reduced GFR, nephrocalcinosis, or deficient AVP, contributes to the presence of a renal concentrating defect in DHT-induced chronic HC.
Interaction of captopril and ibuprofen on glomerular and tubular function in humans
Authors: Allon M. , Pasque C. B. , Rodriguez M.
Published: December 2017

Renal hemodynamics and tubular solute and water handling were evaluated in normal subjects during water diuresis, before and after the acute administration of captopril, ibuprofen, or the combination of both drugs. The glomerular filtration increased after captopril administration but did not change after ibuprofen alone or in combination with captopril. Renal plasma flow increased with captopril alone and captopril plus ibuprofen but did not change after ibuprofen alone. Urine volume and Na excretion increased with captopril and decreased after ibuprofen; coadministration of ibuprofen attenuated the tubular effects produced by captopril alone. FELi, fractional delivery of solute to the distal nephron, and FELi-FENa, fractional distal reabsorption of solute, both significantly increased after captopril and decreased after ibuprofen but did not change with the combined regimen. (FELi-FENa)/FELi, fractional reabsorption of distally delivered Na, significantly decreased after captopril and increased after ibuprofen but remained unchanged after captopril plus ibuprofen. Thus captopril and ibuprofen have opposing effects on tubular Na and water handling, which are attenuated by the addition of the other drug. This interaction may have clinical relevance in patients with heart failure or hypertension, in whom captopril is used to enhance Na and water diuresis.
Role of microfilaments in maintenance of proximal tubule structural and functional integrity
Authors: Kellerman P. S. , Clark R. A. , Hoilien C. A. , Linas S. L. , Molitoris B. A.
Published: December 2017

To determine the selective effect of microfilament disruption on both cellular structure and function, microfilament-specific doses of cytochalasin D (10 microM) were used in an isolated perfused kidney system. Structurally, cytochalasin D resulted in extensive disruption of the apical surface with blebbing, vacuolization, and patchy loss and fusion of microvilli. Functionally, cytochalasin D resulted in an initial decrease in glomerular filtration rate (300.8 +/- 29.9 vs. 541.6 +/- 51 microliters.min-1.g-1, P less than 0.05) with subsequent stabilization throughout the duration of the perfusion. In contrast, the tubular reabsorption of sodium decreased significantly in a linear fashion from 97.1 +/- 0.7 to 64.3 +/- 7.0% over the duration of the perfusion. Similarly, the tubular reabsorption of lithium decreased linearly from 74.8 +/- 2.6%, before the addition of cytochalasin, to 33.6 +/- 6.8% by the end of the perfusion. Correlation of the decrements in percent tubular reabsorption of sodium and lithium for individual kidneys was 0.87 (P less than 0.01), suggesting the effect of microfilament disruption on tubular reabsorption of sodium was localized primarily to the proximal tubule. Because ischemic injury is characterized by time-dependent structural alterations in the apical membrane of proximal tubule cells, we set out to determine whether microfilament disruption occurs during ischemic acute renal failure. Utilizing indirect immunofluorescence with an anti-actin antibody, control kidneys demonstrated intact circumferential apical immunofluorescence representing brush-border and terminal web actin staining. Fifteen minutes of ischemia resulted in multiple large gaps in the terminal web, and 50 min of ischemia caused diffuse redistribution of actin immunofluorescence throughout the cytoplasm.(ABSTRACT TRUNCATED AT 250 WORDS)
Sulfate inhibits [14C]phosphonoformic acid binding to renal brush-border membranes
Authors: Tenenhouse H. S. , Lee J.
Published: December 2017

To examine the specificity of the phosphonoformic acid (PFA) interaction with the Na(+)-dependent phosphate transporter of mouse renal brush-border membrane vesicles, we compared the effects of anions on Na(+)-dependent [14C]PFA binding and Na(+)-dependent phosphate transport. Inhibition of PFA binding was achieved by PFA (% control = 0 +/- 1), sulfate (15 +/- 2), arsenate (35 +/- 1), phosphate (59 +/- 2), and nitrate (68 +/- 4), whereas inhibition of phosphate transport was only apparent with phosphate (0 +/- 1), PFA (22 +/- 4), and arsenate (37 +/- 5). Succinate and gluconate had no effect on either Na(+)-dependent process. Under conditions where Na(+)-dependent PFA binding was maximally inhibited by phosphate (% control = 65 +/- 4), further inhibition could be achieved by sulfate (26 +/- 5%). Na(+)-dependent PFA binding was competitively inhibited by phosphate (apparent Ki = 8.9 +/- 1.2 mM) and noncompetitively inhibited by sulfate (apparent Ki = 2.6 +/- 0.5 mM). We found that PFA inhibited Na(+)-dependent sulfate transport (50% inhibition at 9 mM PFA) as well as Na(+)-dependent phosphate transport (50% inhibition at 0.5 mM PFA). We also examined the pH dependence of Na(+)-dependent PFA binding and Na(+)-dependent phosphate and sulfate transport. PFA binding was optimal at pH = 7.4, whereas phosphate transport increased with increasing pH, and sulfate transport increased with decreasing pH.(ABSTRACT TRUNCATED AT 250 WORDS)
Hypotonic shock activates a maxi K+ channel in primary cultured proximal tubule cells
Authors: Dube L. , Parent L. , Sauve R.
Published: December 2017

The nature and function of the ionic channels at the apical membrane of primary cultured proximal tubule cells (PT) was investigated by use of the extracellular patch-clamp method. Several types of ionic channels were observed, including a calcium-dependent K+ channel of 206 pS in symmetrical 162 mM KCl activated at depolarizing potentials [maxi K+(Ca2+)]. Whole cell experiments were also carried out that clearly indicated that the PT cells respond to a hypotonic shock by activating electroconductive pathways. This response consisted of an initial hyperpolarization (from -47 to -58 mV, SD = 3, n = 4), followed by a strong depolarization (to -23 mV, SD = 4, n = 4). Furthermore, it was found in cell-attached experiments that the maxi K+(Ca2+) channel becomes activated during the hypotonic challenge. The activation process required external Ca2+, although some residual single-channel activity was measured in the absence of extracellular calcium (n = 3). On the basis of these results, it is concluded that the volume regulation process in PT cells in response to a hypotonic shock involves an influx of calcium from the external medium, which in turn triggers the opening of apical maxi K+(Ca2+) channels.
Effects of PTH, calcitonin, and cAMP on calcium transport in rabbit distal nephron segments
Authors: Shimizu T. , Yoshitomi K. , Nakamura M. , Imai M.
Published: December 2017

Distal nephron segments are heterogenous with respect to adenylate cyclase responses to stimulation with parathyroid hormone (PTH) or calcitonin (CT). We examined effects of these hormones and of 8-(p-chlorophenylthio)-adenosine 3',5'-cyclic monophosphate (CPTcAMP) on net Ca absorption (Jnet Ca2+, pmol.min-1.mm-1) in rabbit distal nephron segments by in vitro microperfusion technique. We studied three segments, including distal convoluted tubule (DCT), connecting tubule (CNT), and cortical collecting duct (CCD). PTH (1 nM) in bath significantly increased Jnet Ca2+ from 2.28 +/- 0.35 to 9.44 +/- 1.13 in CNT, but did not affect Jnet Ca2+ in DCT or CCD. CT (1 nM) in bath significantly increased Jnet Ca2+ from 1.58 +/- 0.29 to 4.45 +/- 1.01 in DCT, whereas it did not affect Jnet Ca2+ either in CNT or in CCD. CPTcAMP (30 microM) in bath significantly increased Jnet Ca2+ from 2.29 +/- 0.42 to 3.97 +/- 0.43 in DCT and from 2.43 +/- 0.18 to 5.83 +/- 0.37 in CNT, but it did not affect Jnet Ca2+ in CCD. When Na+ was removed from bathing fluid or when 0.1 mM ouabain was added to bath, Jnet Ca2+ in both DCT and CNT significantly decreased. Furthermore, stimulatory effects of PTH and CT on Ca2+ absorption in the respective segments were abolished under these conditions. These results suggest that PTH and CT increase Ca2+ absorption in CNT and DCT, respectively, through cAMP-mediated mechanisms. Presence of a basolateral Na(+)-Ca2+ exchange process seems to be a prerequisite for effects of these hormones. However, exact intracellular mechanisms remain uncertain.
Phosphorus restriction reverses hyperparathyroidism in uremia independent of changes in calcium and calcitriol
Authors: Lopez-Hilker S. , Dusso A. S. , Rapp N. S. , Martin K. J. , Slatopolsky E.
Published: December 2017

Phosphorus is a well-known modulator of renal 1 alpha-hydroxylase activity. In early and moderate renal failure it is proposed that dietary Pi reduction ameliorates secondary hyperparathyroidism through increased circulating levels of calcitriol (i.e, 1 alpha, 25-dihydroxycholecalciferol). To gain further insight into the mechanisms by which a low-Pi diet ameliorates secondary hyperparathyroidism in advanced renal insufficiency, studies were performed in five dogs before and 6 mo after the induction of uremia by 5/6 nephrectomy. Glomerular filtration rate decreased from 69.0 +/- 2.3 to 10.5 +/- 0.5 ml/min, immunoreactive parathyroid hormone (irPTH) increased from 66.0 +/- 8.8 to 321.0 +/- 46 pg/ml, and calcitriol decreased from 39.0 +/- 10.4 to 27.0 +/- 6.2 pg/ml. Thereafter, dietary Pi was decreased gradually every 2 wk from 0.95% to 0.6, 0.45, and 0.3%, respectively. Dietary Ca was reduced from 1.6 to 0.6% to prevent development of hypercalcemia. Ionized Ca (ICa) decreased from 5.4 +/- 0.04 to 5.2 +/- 0.05 mg/dl (P less than 0.02), and plasma Pi decreased from 6.3 +/- 0.7 to 4.7 +/- 0.2 mg/dl (P less than 0.05). Calcitriol remained low (23.3 +/- 4.7 pg/ml). However, irPTH gradually decreased from 321.0 +/- 46.0 to 94.7 +/- 22.9 pg/ml (P less than 0.005). These studies indicate that a decrease in dietary Pi from 0.95 to 0.3% suppressed irPTH by approximately 70%. Reduction of irPTH was observed in the absence of a concomitant increase in levels of ICa or calcitriol. These studies suggest that reduction in dietary Pi in advanced renal insufficiency improves secondary hyperparathyroidism by a mechanism that is independent of the levels of calcitriol or plasma ICa.
Differential Na fluxes with artificial and native fluids in clip models of volume expansion
Authors: Reddy S. , Cochineas C. , Gyory A. Z.
Published: December 2017

Earlier we demonstrated significant differences in proximal tubular Na transport inhibition during saline volume expansion (VE) depending on whether artificial (AF) or native harvested tubular fluid (HTF) was used. In the present experiments the shrinking-drop technique was used to measure volume flux (Jv) in the early- and late-clip models of VE with AF alternating with HTF. In early-clip rats, with AF, Jv (x 10(4) mm3.mm-2.s-1) during the nonexpanded period was 2.92 +/- 0.105; during subsequent VE, it was 2.20 +/- 0.180 and 1.97 +/- 0.149 with HTF (for the latter two P greater than 0.3). In late-clip rats they were 2.83 +/- 0.135, 1.99 +/- 0.157, and 1.50 +/- 0.171 (for the latter two P less than 0.001), respectively. With AF, transport was inhibited equally in both models during VE. There were no significant differences in the electrolyte composition of AF and HTF during shrinkage as measured by microprobe analysis. The 47% inhibition of Jv with HTF but not AF during VE in the late-clip but not the early-clip rats strongly implies the presence of factors other than physical forces during VE, which inhibit proximal tubular Na transport. Studies of tubular transport during VE need to be performed using both AF and HTF.
Regulation of small-conductance K+ channel in apical membrane of rat cortical collecting tubule
Authors: Wang W. H. , Schwab A. , Giebisch G.
Published: December 2017

We used the patch-clamp technique to study the activity and regulation of single potassium channels in the apical membrane of isolated cortical collecting tubules (CCT) of rat kidney. With 140 mM K+ in the pipette the inward conductance of the channel in cell-attached patches at 37 degrees C was 35 pS (n = 106, NaCl-Ringer or 70 mM KCl and 70 mM NaCl in the bath), and the outward conductance was 15 pS (n = 15, 70 mM NaCl + 70 mM KCl in the bath). Mean open probability (Po) of the channel is voltage independent and 0.96 (n = 106). The channel displayed one open state with a mean lifetime of 18.6 ms and one closed state with a mean lifetime of 0.7 ms (n = 20). Selectivity ratio between K+ and Na+ is 20 (n = 5). High-potassium diet increased channel incidence from control 32% (53 patches with channel from 165 patches) to 64% (53 patches with channels from 83 patches). The channel could be blocked by 1 mM Ba2+ (n = 7, Ba2+ in the pipette); however, 5 mM tetraethylammonium (n = 9, TEA in the pipette) did not block the channel activity. The channel was very sensitive to intracellular pH (n = 6). Changing bath pH facing cytoplasmic side of inside-out patches from 7.4 to 6.9 reversibly reduced Po from 0.9 to 0.1. Addition of 1 mM ATP (n = 7) to bath almost completely inhibited channel activity in inside-out patches. This ATP-induced inhibition was fully reversible and was found to be dependent on the ratio of ATP to ADP, since adding 0.5 mM ADP to bath solution relieved the ATP-induced blockade. Results indicate that intracellular pH, concentration of ATP, and ratio of ATP to ADP are important regulators of potassium channel activity in the apical membrane of rat CCT, and the properties of the channel make it a strong candidate for K+ secretion in this nephron segment.
Glomerular response to hyperglycemia in human diabetic nephropathy
Authors: Remuzzi A. , Viberti G. , Ruggenenti P. , Battaglia C. , Pagni R. , Remuzzi G.
Published: December 2017

We have studied the effect of acute hyperglycemia on glomerular function in seven insulin-dependent diabetics with overt nephropathy during hyperglycemic or euglycemic clamp. In all patients glomerular filtration rate (GFR) was higher during hyperglycemia than during euglycemia (35.0 +/- 15.5 vs. 21.4 +/- 10.3 ml.min-1.1.73 m-2, P less than 0.01), whereas renal plasma flow did not change significantly. To establish which determinant of GFR is altered by hyperglycemia, fractional clearances of neutral dextrans of graded molecular size were determined in both glycemic states, and data were analyzed by a theoretical model of hindered transport of macromolecules through a porous membrane. Hyperglycemia significantly increased sieving coefficients of small dextran molecules (28-40 A in radius), whereas fractional clearances of large macromolecules (greater than 44 A) did not change. Theoretical analysis suggested that the ultrafiltration coefficient (Kf) and membrane permeability to small dextrans (less than 40 A) increased in hyperglycemia in respect to euglycemia. Because these results have been obtained in patients with severe renal failure and hypertension, our conclusions do not necessarily apply to the early phase of diabetic nephropathy.
Na(+)-H+ exchange is stimulated by protein kinase C activation in inner medullary collecting duct cells
Authors: Slotki I. N. , Schwartz J. H. , Alexander E. A.
Published: December 2017

In this study we investigated the role of protein kinases in activation of the Na(+)-H+ exchanger in inner medullary collecting duct (IMCD) cells. Monolayers, 24-48 h after achieving confluence, were made quiescent by 24 h incubation in 0.1% serum before study. Changes in pHi were measured with 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. Phorbol myristate acetate (PMA), a synthetic analogue of diacylglycerol (DAG), was used to stimulate protein kinase C (PKC). In nominally HCO3(-)-free media containing 110 mM Na+ and 1 mM Ca2+, PMA addition increased pHi from 7.29 +/- 0.08 to 7.54 +/- 0.07 after 20 min. The increment in pHi was completely inhibited by 1 mM amiloride or by replacement of extracellular Na+ with choline but not inhibited by 1 mM N-ethylmaleimide, an inhibitor of active proton transport. Downregulation of PKC by overnight incubation of monolayers with PMA also prevented the rise in pHi upon subsequent challenge with PMA. Another active analogue of DAG, 1,2-dioleoyl-rac-glycerol, caused an increment in pHi similar to that produced by PMA, whereas 4 alpha-phorbol, an inactive analogue, did not stimulate Na(+)-H+ exchange. Bradykinin (10(-6) M), a phospholipase C-activating hormone, also induces alkalinization of IMCD cells similar to that produced by phorbol esters. Neither vasopressin (10(-7) M), which induces cellular accumulation of adenosine 3-,5--cyclic monophosphate (cAMP) and activation of protein kinase A (PKA), nor 8-bromo-cAMP (1 mM) changed pHi. Therefore in the IMCD cell activation of PKC but not PKA stimulates a rise in pHi via the Na(+)-H+ exchanger.
High glucose induces cell hypertrophy and stimulates collagen gene transcription in proximal tubule
Authors: Ziyadeh F. N. , Snipes E. R. , Watanabe M. , Alvarez R. J. , Goldfarb S. , Haverty T. P.
Published: December 2017

Tubulointerstitial changes in the diabetic kidney correlate closely with the decline in glomerular filtration. In this study, we used a cell culture system of mouse proximal tubule epithelial cells to test the effects of glucose on cell growth, size, and matrix biosynthesis. [3H]thymidine incorporation was significantly inhibited in cells grown in 450 mg/dl glucose, compared with cells grown in 100 mg/dl glucose. The cells grown in the higher glucose concentration were slightly larger, their protein content and the total protein synthetic rate were significantly increased, and they secreted approximately twice as much procollagens type IV and type I. Concordantly, steady-state procollagen mRNA levels were also increased: 2.6-fold for the alpha 1(IV) and 2.2-fold for the alpha 2(I) procollagens. Additionally, nuclear run-off studies demonstrated that procollagen gene transcription rate was stimulated approximately 50%; beta-actin transcription rate was not altered. We used chloramphenicol acetyltransferase (CAT) reporter gene constructs to determine whether the increased transcription rate of alpha 2(I) gene was associated with activation of its enhancer sequence. Cells transfected with the enhancer demonstrated more than fivefold increase in CAT activity when cultured in the high-glucose medium. These studies demonstrate a multitude of effects of high ambient glucose concentrations on proximal tubule cell growth and collagen biosynthesis; cell proliferation is decreased although cell hypertrophy occurs. Procollagen gene transcription rate is stimulated and this response contributes to the observed increase in procollagen mRNA content. Activation of an enhancer sequence may be one possible mode through which high glucose levels increase the transcription of procollagen type I, presumably involving trans-acting factor(s).