bluuebunny/crossref_metadata_2025 huggingface dataset:
Atrial natriuretic peptide regulates release of Na+-K+-ATPase inhibitor from rat brain
Authors: Crabos M. , Ausiello D. A. , Haupert G. T. , Cantiello H. F.
Published: December 2017

Tissue culture media from incubations of fragments of rat brain were collected and partially purified. These supernatants were effective in inhibiting the Na+-K+ pump as indicated by a 77% reduction of ouabain-sensitive 86Rb+ uptake into human erythrocytes. Release of the Na+-K+-ATPase inhibitor depended on the amount of tissue, the temperature, and the length of incubation. Atrial natriuretic peptide (ANP) injected intravenously, or included (10(-8) M) in the in vitro incubation of brain tissue, decreased the release of the Na+-K+-ATPase inhibitor by 74 and 42%, respectively. Control experiments using the neuropeptide arginine vasopressin showed no effect on release of the inhibitor. These studies indicate that ANP is capable of regulating the release from brain of a Na+-K+-ATPase inhibitor with similar chromatographic characteristics to the one previously obtained from extraction of bovine hypothalamus and raise the possibility that the two factors are interrelated in the regulation of fluid and electrolyte balance.
Renal cytochrome P-450-related arachidonate metabolism in rabbit aortic coarctation
Authors: Carroll M. A. , Schwartzman M. , Baba M. , Miller M. J. , McGiff J. C.
Published: December 2017

Cells of the medullary segment of the thick ascending limb of Henle's loop (TALH) convert arachidonic acid (AA) via the cytochrome P-450 monooxygenase pathway to biologically active metabolites: P1, a vasorelaxant, and P2, an inhibitor of Na+-K+-ATPase activity. These AA metabolites may contribute to the renal vascular and metabolic adjustments in response to renal hypoperfusion and the attendant elevation of blood pressure produced by suprarenal aortic coarctation. On the eighth postoperative day, the blood pressures of hypertensive and sham-operated control rabbits were 105 (90-115) and 63 (60-64) mmHg (medians with semiquartile values), respectively (P less than 0.01). Formation of P1 and P2 was increased twofold in TALH cells obtained from hypertensive rabbits: 2.35 (1.79-4.83) and 1.28 (1.56-4.56) micrograms AA converted.mg protein-1.30 min-1 compared with sham-operated rabbits: 1.27 (1.03-1.53) and 0.64 (0.58-1.10) micrograms AA converted.mg protein-1.30 min-1 (P less than 0.05). The profile of biological activity of AA metabolites contained within P1 and P2 was unaffected by aortic coarctation. The cytochrome P-450 monooxygenase-derived AA metabolites may exert a defensive function to limit the degree of TALH cell injury in response to renal hypoperfusion and associated zonal anoxia by reducing energy-dependent Na+-K+-ATPase activity and affecting local vasodilatation.
Electrophysiological properties of amphibian late distal tubule in vivo
Authors: Planelles G. , Anagnostopoulos T.
Published: December 2017

This study was undertaken to determine the passive electrophysiological properties of the diffusive barriers of the late distal tubule (LDT) in Necturus. The transepithelial resistance (RT) determined by cable analysis was 1,130 omega.cm2, which puts the LDT in the class of "tight" epithelia. Using two different methods, we did not find significant cell-to-cell electrical coupling. The fractional apical resistance was 0.93, and it did not vary with distance from the current-injecting electrode. Relative permeabilities of K+, Na+, and Cl- during peritubular ion concentration changes were assessed by circuit analysis. The conclusions are as follows. The basolateral cell membrane is highly permeable to K+; its apparent K+ transference number is 0.78. Basolateral chloride transference was very small. Sodium removal from peritubular fluid produced depolarization, suggesting carrier-mediated electrogenic Na+ transport. The high fractional resistance of the apical cell membrane prevented assessment of apical transference numbers. However, Cl- removal from luminal fluid produced cell hyperpolarization; the underlying mechanism has not been established with certainty. The paracellular pathway does not discriminate between Na+, Cl-, and some of their substitutes; it is poorly permeable to gluconate and prefers K+ to Na+.
Complementary role of citrate and bicarbonate excretion in acid-base balance in the rat
Authors: Kaufman A. M. , Kahn T.
Published: December 2017

Studies were performed to evaluate whether alterations in the excretion of citrate, a metabolic precursor of bicarbonate, play a quantitatively important role in acid-base balance during bicarbonate feeding in the rat. Potassium depletion (K-DEPL), chloride depletion (Cl-DEPL), or potassium plus chloride depletion (KCl-DEPL) was produced by eliminating potassium, chloride, or potassium chloride from the diet. After 3 days of depletion, sodium bicarbonate (4,000 mueq/24 h) was added to the diet for 7 days. In all groups plasma bicarbonate concentration increased minimally during bicarbonate administration and was similar to normal controls receiving bicarbonate. In K-DEPL, citrate excretion was less than normal but bicarbonate excretion was greater than normal. In Cl-DEPL, bicarbonate excretion was less than normal but citrate excretion was greater than normal. In KCl-DEPL, bicarbonate and citrate excretion were similar to normal. Sodium bicarbonate was also administered to K-DEPL and KCl-DEPL rats in which plasma bicarbonate concentration averaged 32.9 meq/1. The reciprocal relationship between citrate and bicarbonate excretion was not altered by the profound metabolic alkalosis. Again, plasma bicarbonate concentration changed little with sodium bicarbonate administration. These studies suggest that the ability to excrete a base load remains intact despite potassium or chloride depletion or metabolic alkalosis. Complementary alterations of citrate and bicarbonate excretion play an important role in acid-base balance under these conditions.
Innervation of the thick ascending limb of Henle
Authors: Barajas L. , Powers K. V.
Published: December 2017

The overlap of accumulations of autoradiographic grains (AAGs) on profiles of the thick ascending limb of Henle (TALH) was measured in autoradiograms of sections from rat kidneys with monoaminergic nerves labeled by means of tritiated norepinephrine. The amount of AAG overlap was used as an indirect means of quantifying innervation along the TALHs of superficial, mid-cortical, and juxtamedullary nephrons. The density of innervation along the TALH showed nephron heterogeneity; the juxtamedullary nephrons with a high pre- and postjuxtaglomerular apparatus (JGA) TALH density of innervation and the upper and midcortical nephrons with high TALH innervation densities at the level of the JGA. The pre-JGA TALH of the juxtamedullary nephrons had a significantly higher (P less than 0.001) density of innervation than the midcortical or superficial nephrons. The TALHs of juxtamedullary nephrons were found to have substantially more innervation than the TALHs of the other nephrons. For all three populations of nephrons, the pre-JGA TALH had the greatest amount of innervation. Neural regulation of TALH function would occur mainly along the pre-JGA and level of the JGA TALH. This regulation would increase TALH NaCl reabsorption (decrease luminal NaCl concentration) and therefore influence 1) the urinary concentrating mechanism, and 2) renin secretion via the macula densa mechanism. The innervation of the TALH was predominantly associated with the vasculature of the TALH's own nephron. However, innervation associated with medullary ray capillary beds from deeper nephrons was observed on pre-JGA TALHs from superficial and midcortical nephrons.
Characterization of prazosin-sensitive alpha 2 B-adrenoceptors expressed by cultured rat IMCD cells
Authors: Yasuda G. , Sun L. , Umemura S. , Pettinger W. A. , Jeffries W. B.
Published: December 2017

alpha 2-Adrenoceptor subtype expression was investigated in cultured rat inner medullary collecting duct (IMCD) cells using radioligand binding studies, Northern blot analysis, and adenosine 3',5'-cyclic monophosphate (cAMP) assays. [3H]rauwolscine bound to a single class of alpha 2-adrenoceptors with high affinity [Kd = 1.7 +/- 0.3 nM, maximum binding (Bmax) = 45.2 +/- 10.8 fmol/mg protein]. alpha 2-Adrenoceptor ligands inhibited [3H]rauwolscine binding with a rank order of potency characteristic of interaction with the alpha 2B-adrenoceptor [inhibitory constant (Ki) values (in nM) rauwolscine (1.95) greater than ARC-239 (8.52) greater than prazosin (237) greater than oxymetazoline (30,000)]. Northern blot analysis was performed using poly(A)+ RNA isolated from 90% confluent rat IMCD cells and probes derived from alpha 2-adrenoceptor DNA sequences from the rat nonglycosylated alpha 2B-adrenoceptor and the human alpha 2A-adrenoceptor. The alpha 2B probe hybridized to a 4.2-kb band under high stringency conditions, but the alpha 2A-adrenoceptor probe did not hybridize to this band. In functional studies, the full alpha 2-adrenoceptor agonists epinephrine and UK-14,304 potently inhibited vasopressin-stimulated cAMP accumulation by 50 to 70% [half-maximal response (EC50) (in nM) epinephrine = 11.2, UK-14,304 = 6.4]. Guanabenz and clonidine were partial agonists, inhibiting cAMP accumulation by 30 to 40% and were less potent than the full agonists [EC50 (in nM) 56.0 guanabenz and 94.5 clonidine]. Epinephrine-induced inhibition of cAMP accumulation was blocked by rauwolscine, prazosin, and ARC-239 but not by the alpha 1-adrenoceptor antagonist corynanthine. We conclude that rat IMCD cells in primary culture express functional alpha 2-adrenoceptors of the alpha 2B-subtype.
Evidence that nephrocalcin and urine inhibit nucleation of calcium oxalate monohydrate crystals
Authors: Asplin J. , DeGanello S. , Nakagawa Y. N. , Coe F. L.
Published: December 2017

Human urine, and nephrocalcin (NC), a glycoprotein of probable kidney cell origin, greatly reduce consumption of calcium and oxalate from metastably supersaturated solutions seeded with calcium oxalate crystals, a phenomenon usually referred to as inhibition of crystal growth. We seeded metastably supersaturated calcium oxalate solutions with calcium oxalate monohydrate crystals under conditions of ion clamping to maintain constant composition and measured ion consumption from pump delivery rates. Consumption rates increased continuously with time as if the solutions were autocatalytic. After incubation, the seeds were covered with innumerable crystallites, which were also free and numerous in the solution, reflecting self-nucleation. The addition of 20% whole, dialyzed urine, or purified NC reduced ion consumption rates markedly, and the only crystals observed at the end of incubation were the large original seeds. Crystals precoated with concentrated dialyzed urine or NC also showed reduced ion consumption. Urine and NC from patients with nephrolithiasis inhibited nucleation less than normal controls. Self-nucleation seems to be the preferred response in sparsely seeded, ion-clamped, supersaturated solutions, such as exist in the nephron. Urine and NC suppress self-nucleation in vitro by adsorbing to the surface of calcium oxalate crystals.
Heterogeneous localization of G protein alpha-subunits in rat kidney
Authors: Stow J. L. , Sabolic I. , Brown D.
Published: December 2017

The Gs alpha and Gi alpha 1-3 subunits of GTP-binding proteins were localized in sections of rat kidney using antibodies against unique synthetic decapeptides from the different G alpha subunits. All of the G alpha subunits were found to have a polarized distribution on renal tubule epithelial cells, and staining was typically found on either basolateral or apical membranes in a given cell type. Gi alpha 1 was localized to the apical pole of both thick ascending limb cells and cells forming the papillary epithelium, Gi alpha 2 labeled the basolateral plasma membrane and the cytoplasm of collecting duct principal cells, and Gi alpha 3 was most abundant in the apical region of proximal tubule cells of the S1 segment, where it was concentrated in sub-brush-border invaginations. It was also found in the perinuclear Golgi complex in these cells. Gs alpha was heavily concentrated on the basolateral plasma membranes of thick ascending limb cells and both principal and intercalated cells of the collecting duct. Less intense subapical staining of G alpha s was also found in proximal tubule cells. The cells of the macula densa had a unique G protein distribution that was distinct from the surrounding cells of the thick ascending limb of Henle. Antibodies specific for the Gi alpha 1 and Gi alpha 3 subunits both stained intracellular vesicles clustered at the basal pole of the cell. A heterogeneous distribution of G alpha subunits was also found by Western blotting on isolated cortical membrane fractions.
DA1 dopamine receptors in renal cortical collecting duct
Authors: Ohbu K. , Felder R. A.
Published: December 2017

Renal dopamine DA1 receptors are linked to the regulation of sodium transport. We have previously reported the presence of DA1 receptors in the proximal convoluted tubule (PCT) but not in the distal convoluted tubule. However, the DA1 receptor in the collecting duct, the final determinant of electrolyte transport, has not been studied. DA1 receptors were studied in the microdissected cortical collecting duct (CCD) of rats by autoradiography with use of the selective DA1 radioligand 125I-Sch 23982 and by measurement of adenylate cyclase (AC) activity. Specific binding of 125I-Sch 23982 to CCD was saturable with radioligand concentration. The dissociation constant (Kd) was 0.46 +/- 0.08 nM (n = 5), and the maximum receptor density (Bmax) was 1.41 +/- 0.43 fmol/mg protein (n = 5). The DA1 antagonist Sch 23390 was more effective than the DA1 agonist fenoldopam in competing for specific 125I-Sch 23982 binding. Fenoldopam stimulated AC activity in CCD in a concentration-dependent (10(-9)-10(-6) M) manner. The ability of fenoldopam to stimulate AC activity was similar in CCD and PCT even though DA1 receptor density was 1,000 times greater in the CCD than in the PCT. In additional studies, fenoldopam stimulation of AC activity did not influence vasopressin-stimulated AC activity. We conclude that the DA1 receptor in rat CCD is tightly coupled to AC stimulation and that there is no interaction between DA1 agonist-stimulated and vasopressin-stimulated AC activity in the CCD.
Mechanism of sodium transport inhibition by epidermal growth factor in cortical collecting ducts
Authors: Vehaskari V. M. , Herndon J. , Hamm L. L.
Published: December 2017

Epidermal growth factor (EGF) inhibits Na transport in the cortical collecting ducts (CCD). To gain insight into the signal transduction of this effect, several potential mechanisms were examined in rabbit CCD perfused in vitro. Pretreatment with pertussis toxin, indomethacin, or the protein kinase C inhibitor H7 did not prevent the acute 34-50% decrease in lumen-to-bath 22Na flux (JNa) on exposure to peritubular EGF, indicating that the inhibition is not mediated by a Gi protein, prostaglandin E2 (PGE2), or protein kinase C. Inhibition of the basolateral Na-H exchanger was also without an effect. Lowering the bath Ca concentration from 1.2 to 0.11 mM did not prevent the inhibition of JNa by EGF (JNa decreased significantly by 38.7 +/- 6.9% and 29.1 +/- 5.3%, respectively); in contrast, reduction of the bath free Ca to 0.005 mM totally abolished the effect of EGF. The response to EGF was also assessed in the setting of chronic stimulation of Na transport; inhibition of JNa by EGF was still observed in CCD from remnant kidneys and in CCD from mineralocorticoid-treated rabbits. The results demonstrate that the inhibition of CCD Na transport by EGF is dependent on peritubular Ca. This suggests that the signal transduction involves Ca influx across the basolateral membrane and that increased cytosolic free Ca may be a common pathway for the counterregulatory control of Na reabsorption by several agonists.
Renal natriuretic peptide (urodilatin?) and atriopeptin: evolving concepts
Authors: Goetz K. L.
Published: December 2017

The discovery of the natriuretic peptide, urodilatin, in human urine, together with data from renal cell cultures, immunocytochemistry, radioimmunoassays, and other techniques, strongly indicates that distal tubules in the kidney produce a natriuretic prohormone that may be identical to the 126-residue prohormone of atrial natriuretic peptide [proANP-(1-126)] produced by cardiac atria. The 32-residue peptide, urodilatin [ANP-(95-126)], is found in urine but not in plasma; its natriuretic potency equals or exceeds that of atriopeptin [ANP-(99-126)]. It is still unclear whether urodilatin is the only, or even the primary, renal natriuretic peptide. Circumstantial evidence suggests that natriuretic peptide produced in connecting and collecting tubules may be a principal physiological ligand for more distal "ANP" receptors, but this issue is far from settled. Evidence that the kidneys produce their own natriuretic peptide may partly explain why investigations of the effects of endogenous atriopeptin on renal sodium excretion often have yielded inconsistent results. This inconsistency has led to a gradual evolution of opinion concerning the functional role of atriopeptin. Based on current data, it is postulated that atriopeptin is primarily a regulator of the cardiovascular system and that renal natriuretic peptide participates in the intrarenal regulation of sodium and chloride transport.
Osmotically regulated taurine content in rat renal inner medulla
Authors: Nakanishi T. , Uyama O. , Sugita M.
Published: December 2017

During antidiuresis renal medullary cells were previously found to accumulate large amounts of organic osmolytes, namely sorbitol, myo-inositol, glycerophosphorylcholine (GPC), and betaine. Large quantities of amino acids are also present in the renal medulla, but it has been questionable whether the levels of medullary amino acids are osmotically regulated. Therefore we directly measured 26 different amino acids, as well as sorbitol, myo-inositol, GPC, and betaine alone the corticomedullary axis of rats that were either salt loaded, after infusion of hypertonic NaCl solution, or were hydrated, after infusion of hypotonic NaCl solution. The amounts of sorbitol, myo-inositol, GPC, and betaine are greater in the inner medullas of salt-loaded rats compared with hydrated rats. In addition, the amount of taurine is much greater in the inner medullas of salt-loaded rats. Aspartic acid also increases in salt-loaded rats but to a lesser extent. There are substantial gradients of taurine, aspartic acid, and some of 24 other measured amino acids along the corticomedullary axis. However, taurine and aspartic acid are the only measured amino acids that increase significantly during salt loading.
Kinetic analysis of clearance of epidermal growth factor in isolated perfused rat kidney
Authors: Kim D. C. , Hanano M. , Sawada Y. , Iga T. , Sugiyama Y.
Published: December 2017

Our previous in vivo studies identified the saturable uptake of epidermal growth factor (EGF) by rat kidney (D.C. Kim, Y. Sugiyama, H. Sato, T. Fuwa, T. Iga, and M. Hanano, J. Pharm. Sci. 77: 200-207, 1988). In the present study, renal handling of EGF in filtering and nonfiltering isolated perfused rat kidneys was investigated. At designated times after the recirculatory perfusion of 125I-EGF in the nonfiltering kidney, the surface-bound and internalized EGF were separately determined by an acid-washing technique. Time profiles of cell-surface-bound and internalized EGF obtained at the perfusion of a tracer 125I-EGF were fitted to the pharmacokinetic model, and kinetic parameters obtained were as follows: konRs = 0.49 ml.min-1.g-1, koff = 0.87 min-1, kint = 0.20 min-1 where konRs is the binding clearance of EGF with its receptor and koff and kint represent the dissociation and internalization rate constants of the EGF-receptor complex, respectively. The Scatchard analysis of the concentration-dependent EGF binding in the nonfiltering kidney suggests the presence of two binding components, one with high affinity [the apparent dissociation constant (Kd1 = 0.1 nM) and the other with low affinity (Kd2 = 30 nM]. By comparing the internalization clearance (CLi) with filtering and nonfiltering kidneys, we concluded that the renal uptake of EGF occurs mainly from the antiluminal side via receptor-mediated endocytosis in a saturable manner and that the nonsaturable reabsorption of EGF from the luminal membrane after glomerular filtration is relatively small. The contribution of reabsorption, however, becomes larger with the increase in EGF concentration.
Effects of angiotensin on renal cortical and papillary blood flows measured by laser-Doppler flowmetry
Authors: Nobes M. S. , Harris P. J. , Yamada H. , Mendelsohn F. A.
Published: December 2017

The effects of angiotensin II (ANG II) or angiotensin III (ANG III) on renal cortical blood flow (CBF) or papillary blood flow (PBF) were investigated in Inactin-anesthetized young rats with the use of laser-Doppler flowmetry. Infusion of equimolar pressor doses of ANG II (300 ng.kg-1.min-1 iv) or ANG III (267 ng.kg-1.min-1) decreased CBF by 31 +/- 2.6% (P less than 0.001) and 20.3 +/- 3.2% (P less than 0.01), respectively but increased PBF by 19 +/- 6.1% (P less than 0.05) and 14.6 +/- 4.4% (P less than 0.05). The ANG II-induced increase in PBF was not prevented by aortic clamping to maintain constant renal perfusion pressure or pretreatment with the prostaglandin synthase inhibitor, indomethacin. The nonpeptide ANG II receptor antagonist, DuP 753 completely abolished the systemic and intrarenal effects of ANG II. After pretreatment with a kallikrein inhibitor, aprotinin, ANG II infusion increased mean arterial pressure but did not affect PBF, suggesting that kinins, but not prostaglandins, modulate the action of systemic ANG II on PBF. We conclude that circulating ANG II induces vasoconstriction in the cortex and also promotes the intrarenal production of kinins, which act to enhance papillary blood flow.
Ammonium and bicarbonate transport in rat outer medullary collecting ducts
Authors: Flessner M. F. , Wall S. M. , Knepper M. A.
Published: December 2017

Previous in vitro studies have demonstrated spontaneous bicarbonate absorption in the outer stripe portion of the rat outer medullary collecting duct (OMCD) and inner medullary collecting duct, but net acid transport has not been studied in the inner stripe of the rat OMCD (OMCDIS). When we perfused isolated OMCDIS segments with identical bath and perfusate solutions containing HCO-3 and NH4Cl, HCO-3 was spontaneously absorbed, and total ammonia was spontaneously secreted at rapid rates in tubules from both deoxycorticosterone (DOC)-treated and untreated rats. We next measured the NH3 flux due to imposed NH3 concentration gradients. Carbonic anhydrase (CA), when added to the lumen, enhanced the NH3 flux, implying an absence of endogenous CA. The NH3 permeability was 0.0042 +/- 0.0007 cm/s. By measuring the luminal pH in perfused OMCDIS segments with an imposed lumen-to-bath NH3 gradient, we determined the pH at the end of the lumen to be 0.23 units below the equilibrium pH calculated from the simultaneously measured total CO2 concentration in collected fluid, confirming the lack of luminal CA. These results are consistent with the view that ammonium secretion in the OMCDIS occurs predominantly by H+ secretion and parallel NH3 diffusion. A luminal disequilibrium pH due to H+ secretion in the absence of endogenous luminal CA enhances the NH3 entry rate. Spontaneous net acid secretion appears to occur more rapidly in the OMCD than in other parts of the rat collecting duct system.
Renal vascular reactivity to vasodilator prostaglandins in genetically hypertensive rats
Authors: Chatziantoniou C. , Arendshorst W. J.
Published: December 2017

The objective of this study was to test the hypothesis that the vasodilator prostaglandins E2 (PGE2) and PGI2) participate in the mechanisms involved in the increased renal vascular reactivity (RVR) observed in genetic hypertension. Studies were performed on anesthetized young and adult spontaneously hypertensive (SHR) and Wistar-Kyoto rats (WKY). Renal blood flow (RBF) was measured during bolus injections into the renal artery of different doses of viprostol and iloprost (stable receptor agonists of PGE2 and PGI2, respectively) before and during inhibition of prostaglandin synthesis by indomethacin. Under control conditions, PGE2 increased RBF equally in young SHR and WKY. However, after cyclooxygenase inhibition the PGE2-induced increase in RBF was larger in young SHR than in WKY. Adult SHR displayed reduced reactivity to PGE2 relative to age-matched WKY under control conditions. This strain difference was abolished after indomethacin administration. PGI2 increased RBF slightly in young rats before and after indomethacin administration. In contrast, both strains of older animals displayed significant increases in RBF in response to PGI2 injections. Indomethacin administration enhanced this PGI2-induced relaxation in adult SHR but not WKY. We propose that the action of vasodilator PGs in the renal vasculature of rats developing hypertension may be limited by low density of their renal receptors and/or the opposing action of vasoconstrictor PGs. As age advances, PGI2 seems to be activated, possibly as part of a regulatory response to counterbalance the increased renal vascular resistance following the establishment of the disease.
Role of cytoskeleton in volume regulation of rabbit proximal tubule in dilute medium
Authors: Linshaw M. A. , Fogel C. A. , Downey G. P. , Koo E. W. , Gotlieb A. I.
Published: December 2017

When proximal tubules are immersed in hypotonic medium, they quickly swell to a peak volume. In a second, slower phase, termed volume regulatory decrease (VRD), they shrink as K, anion, and water leave the cells. We investigated the role of the cytoskeleton during this biphasic hypotonic volume regulatory response. Isolated, collapsed rabbit proximal convoluted tubules (PCT) were crimped tightly between two pipettes, and their volume was assessed optically. PCT volume increased to a peak 70-80% above baseline on sudden immersion in dilute medium (150 mosmol/kgH2O). After completing VRD, control tubules had regulated their volume 73 +/- 2% back toward baseline. Tubules exposed to the microtubule inhibitor vincristine (5 microM) regulated 75 +/- 2%. Tubules exposed to the microfilament inhibitor cytochalasin B (50 microM) regulated less (57 +/- 5%), and tubules exposed to both inhibitors regulated only 39 +/- 3% (P less than 0.01 vs. control). Hypotonic VRD was unimpaired in PCT loaded with NaCl by prior exposure to ouabain but was significantly reduced by cytochalasin B. We conclude that VRD is not cation specific and that intact microtubules and microfilaments play a synergistic role in the VRD of rabbit PCT in hyposmotic medium.
Assembly of distinctive coated pit and microvillar microdomains in the renal brush border
Authors: Biemesderfer D. , Dekan G. , Aronson P. S. , Farquhar M. G.
Published: December 2017

The plasma membrane of the kidney brush border is composed of two compositionally distinct microdomains, microvilli and clathrin-coated pits. To study their assembly we have immunolocalized brush border marker proteins in the developing proximal tubule epithelium of the neonatal rat and compared their time and site of appearance with those of basolateral markers, Na-K-ATPase and fodrin. The proteins studied were dipeptidyl peptidase IV (DPPIV) (microvilli), actin and villin (microvillar cytoskeletal proteins), glycoprotein 330 (gp330) (coated pits), and clathrin (coated pit cytoskeleton). Although apical microvilli and coated pits were first seen in the stage III nephron, many brush border markers including DPPIV, actin, and clathrin appeared earlier in the development and initially were not polarized. Only during stage III did they become concentrated at the apical membrane. Villin first appeared in the stage III proximal tubule where it was located diffusely in the cytoplasm and in lysosomes as well as along the apical membrane. It did not completely colocalize with actin until stage IV. Gp330 first appeared during stage III and from the beginning was restricted to the apical clathrin-coated membrane domains and endosomes. The results demonstrate that 1) the expression of renal brush border proteins during development is asynchronous, and 2) unlike the basolateral plasmalemmal domain, which is established early in nephrogenesis, brush border assembly occurs later, approximately coinciding with the onset of glomerular filtration.
Regulation of glycolytic metabolism during long-term primary culture of renal proximal tubule cells
Authors: Aleo M. D. , Schnellmann R. G.
Published: December 2017

Adequate oxygenation was a major factor regulating the induction of glycolytic metabolism in primary cultures of rabbit renal proximal tubule cells during short-term (less than 1 day) and long-term (1-7 day) culture. As measured by cellular lactate content, glucose consumption, and lactate dehydrogenase activity, less glycolytic metabolism was induced in cultured cells that were constantly aerated than in cells that were held stationary. When oxidative metabolism is supported by providing 2-10 mM heptanoate (HEP) as a metabolic substrate glycolytic metabolism further decreased during short-term, but not long-term culture. Cellular proliferation did not play a major role in regulating the induction of glycolytic metabolism, since glycolytic metabolism increased before cell growth had occurred, did not decline once logarithmic cell growth had ceased, and was stimulated less by cell growth than by inadequate oxygenation. Fructose-1,6-bisphosphatase and alkaline phosphatase, representative markers of gluconeogenic and brush-border membrane enzyme activities, respectively, declined during culture regardless of culture conditions or the presence of HEP. Therefore, glycolytic metabolism can be effectively minimized by constantly aerating cultured proximal tubule cells and can be further reduced by the addition of HEP during short-term culture.
Response of intercalated cells to chloride depletion metabolic alkalosis
Authors: Verlander J. W. , Madsen K. M. , Galla J. H. , Luke R. G. , Tisher C. C.
Published: December 2017

We examined the effect of Cl- depletion metabolic alkalosis (CDA) on H(+)-ATPase and band 3 protein localization in intercalated cells (IC) of the rat cortical collecting duct (CCD) and the outer medullary collecting duct (OMCD). After 30 min of peritoneal dialysis against 0.15 M NaHCO3 to produce CDA, or Ringer bicarbonate to serve as controls (CON), both groups were infused intravenously with an 80 mM Cl- solution for 90 min. For CDA vs. CON, physiological parameters were as follows: plasma total CO2, 38.0 +/- 1.1 vs. 27.8 +/- 0.6 meq/l (P less than 0.001); urinary total CO2 excretion, 141 +/- 89 vs. 20 +/- 3 neq.min-1.100 g body wt-1; and urinary Cl- excretion, 20 +/- 10 vs. 486 +/- 144 neq.min-1.100 g body wt-1 (P less than 0.001). H(+)-ATPase was localized in thin sections using a rabbit polyclonal antibody against the 70-kDa subunit of bovine brain H(+)-ATPase. Band 3 protein was localized using a polyclonal antibody against the 43-kDa subunit of the cytoplasmic domain of human erythrocyte band 3 protein. In CON rats, H(+)-ATPase localized along the apical plasma membrane and over the apical cytoplasmic vesicles of type A ICs in the CCD and ICs of the OMCD. H(+)-ATPase was observed along the basolateral plasma membrane and over cytoplasmic vesicles throughout type B ICs. In CDA rats, H(+)-ATPase was only observed over apical cytoplasmic vesicles in type A ICs and in the majority of OMCD ICs. In type B ICs, H(+)-ATPase staining was intensified along the basal plasma membrane in CDA. Band 3 protein was consistently localized in the basolateral plasma membrane of all type A cells in the CCD and ICs of the OMCD in both CON and CDA. In summary, stimulation of HCO3- secretion in rats caused withdrawal of H(+)-ATPase from the apical plasma membrane and storage in apical cytoplasmic vesicles of ICs of the OMCD and type A ICs of the CCD. H(+)-ATPase appeared to be inserted into the basal plasma membrane of type B ICs. These findings suggest that, during correction of CDA, proton secretion by type A and OMCD ICs is suppressed and proton transport across the basolateral plasma membrane of type B ICs is stimulated.
Plasma IgG pool is not defended from urinary loss in nephrotic syndrome
Authors: al-Bander H. A. , Martin V. I. , Kaysen G. A.
Published: December 2017

In the nephrotic syndrome, the plasma level of some proteins of hepatic origin is partially defended by an increase in their synthetic rate. The plasma levels of several liver-derived proteins, as well as immunoglobulin G (IgG) are increased in one condition where plasma albumin concentration and oncotic pressure (II) are reduced, i.e., hereditary analbuminemia. To determine whether the urinary loss of IgG, a protein derived from the immune system, is compensated for by an increased synthetic rate, we measured IgG synthesis in normal Sprague-Dawley rats (SD); two models of the nephrotic syndrome: Heymann nephritis (HN) and Adriamycin treatment; and in Nagase analbuminemic rats (NAR), a model of decreased II without urinary protein loss. Plasma IgG and total IgG mass were significantly reduced in both HN and Adriamycin, yet IgG synthesis was nearly identical in HN, Adriamycin, and SD. In contrast, plasma and total IgG and IgG synthesis were all markedly increased in NAR. We derived a pathogen-free colony of NAR by Caesarean section and found that plasma IgG was not increased in pathogen-free NAR, despite reduced II. Thus, unlike proteins of hepatic origin (e.g., albumin) where synthesis increases following urinary loss, no compensatory increase in IgG synthesis occurs. Increased plasma IgG as well as IgG synthesis in the NAR is not a compensatory response to the absence of albumin or reduction in II, but rather is due to subclinical infection. Profound hypogammaglobulinemia of nephrotic syndrome occurs in part because no compensatory synthetic mechanisms balance urinary loss, and alteration in plasma II does not modulate IgG synthesis.
Effect of peritubular hypertonicity on water and urea transport of inner medullary collecting duct
Authors: Kudo L. H. , Cesar K. R. , Ping W. C. , Rocha A. S.
Published: December 2017

The effect of bath fluid hypertonicity on hydraulic conductivity (Lp) and [14C]urea permeability (Pu) of the distal inner medullary collecting duct (IMCD) was studied in the absence and in the presence of vasopressin (VP) using the in vitro microperfusion technique of rat IMCD. In the first three groups of IMCD, we observed that in the absence of VP the Lp was not different from zero when the osmotic gradient was created by hypotonic perfusate and isotonic bath fluid, but it was significantly greater than 1.0 x 10(-6) cm.atm-1.s-1 when the osmotic gradient was created by hypertonic bath and isotonic perfusion fluid. The increase in Lp was observed when the hypertonicity of the bath fluid was produced by the addition of NaCl or raffinose, but no such effect was observed with urea. The stimulated effect of bath fluid hypertonicity on Lp was also observed in the IMCD obtained from Brattleboro homozygous rats in which VP is absent. The NaCl hypertonic bath increased the Pu in the absence of VP. In another series of experiments with VP (10(-10) M) we observed that the hypertonic bath fluid increased in a reversible manner the VP-stimulated Lp of distal IMCD. However, the NaCl hypertonicity of the bath fluid was not able to increase dibutyryladenosine 3',5'-cyclic monophosphate-stimulated Lp. The Pu stimulated by VP (10(-10) M) increased twofold when the bath fluid was hypertonic. Therefore hypertonicity of the peritubular fluid produced by the addition of NaCl or raffinose increases the Lp and Pu in the absence and in the presence of VP. No such effect was noted with the addition of urea.
Morphometric analysis of the actions of angiotensin II on renal arterioles and glomeruli
Authors: Denton K. M. , Fennessy P. A. , Alcorn D. , Anderson W. P.
Published: December 2017

To study the effects of angiotensin II on afferent and efferent arteriole diameters and on intraglomerular dimensions, angiotensin II (20 ng.kg-1.min-1) or saline vehicle was infused intravenously for 20 min into anesthetized rabbits pretreated with enalapril. Both kidneys were perfusion fixed (glutaraldehyde), and vascular casts were made of the right kidneys using methacrylate. Morphometric analysis of the left kidneys using transmission electron microscopy revealed no significant effects of angiotensin II within the glomerulus, including the degree of mesangial contraction. The diameters of the afferent and efferent arteriole casts from the right kidneys were measured at 20, 50, and 75 microns from the glomerulus by scanning electron microscopy. In the outer cortex the mean diameters of the afferent and efferent arterioles were 14.1 +/- 0.8 and 9.7 +/- 0.5 microns, respectively, in the angiotensin II-infused rabbits, significantly less than in the control (vehicle) rabbits, 17.0 +/- 0.7 microns (P less than 0.001) and 10.7 +/- 0.4 microns (P less than 0.005), respectively. Calculation of the relative changes in vascular resistance, however, indicated that the effects of angiotensin II on efferent arteriole resistance (average difference 2.4 +/- 1.2 units/microns) were significantly greater per unit length than the effects on afferent arteriole resistance (average difference 0.9 +/- 0.3 units/microns). Thus infused angiotensin II caused greater reduction in afferent arteriolar diameter than in efferent, but the calculated increase in vascular resistance per micron was greater in efferent vessels due to their smaller resting diameter.
Hypotonicity and cell volume regulation in shark rectal gland: role of organic osmolytes and F-actin
Authors: Ziyadeh F. N. , Mills J. W. , Kleinzeller A.
Published: December 2017

Hypotonic stress (reduction of external tonicity from approximately 900 mosM and 295 mM NaCl to approximately 600 mosM and 135 mM NaCl) produced a relatively slow regulatory volume decrease (RVD) in dogfish shark (Squalus acanthias) rectal gland cells. During the 5-h experiment, cell K+ content remained unchanged; cell content of Na+ and Cl- dropped in the initial swelling phase by some 50% (reflecting the corresponding reduction in medium NaCl), and then remained unchanged during volume recovery phase. Also, cellular fluxes of 86Rb+ and urea were not affected by hypotonic stress. However, hypotonicity enhanced 10- to 20-fold the efflux of organic cell osmolytes taurine, betaine, and trimethyloxamine, and this accounted for the loss of osmotically obliged water during RVD. Enhancement of osmolyte efflux by hypotonic stress was abolished by readjusting the low-Na+ saline to isotonicity (approximately 900 mosM) with innocuous cations (choline+, Li+, or N-methylglucamine+). The results suggest that reduction of medium tonicity may be the determinant for the RVD response to hypotonic stress. The above properties of the observed RVD were also displayed when studying changes on cell F-actin at the basolateral cell face; hypotonic stress (medium with 135 mM NaCl) produced a rapid disappearance of fluorescence related to this cytoskeletal component, whereas no such changes were seen in low-Na+ salines made isotonic with choline or N-methylglucamine chloride nor in a saline made hyposmolar by omitting urea. Hence, hypotonicity is required to affect F-actin organization (depolymerization?). These changes of F-actin fluorescence are transient; they were completed within 5-10 min of hypotonic stress, and afterwards a gradual reconstitution of cell F-actin organization was seen. The above observations are consistent with the assumption that, in shark rectal gland cells, transient loss of cytoskeleton (F-actin) organization at the basolateral cell face, induced by hypotonicity, brings about a selective efflux of organic osmolytes, thus producing the observed RVD.
Dopamine receptor-mediated activation of phospholipase C is associated with natriuresis during high salt intake
Authors: Vyas S. J. , Jadhav A. L. , Eichberg J. , Lokhandwala M. F.
Published: December 2017

Activation of phospholipase C (PLC) is considered to be one of the cellular signaling events involved in dopamine (DA)-mediated natriuresis. In the present study we have examined the role of renal cortical PLC in contributing to the increase in urinary sodium excretion during high sodium intake and its relationship with intrarenal DA synthesis. Rats were given either 1% NaCl (high sodium intake) or tap water (normal sodium intake) to drink for 24 h, and urine was collected over this time period. PLC activity in the renal cortex from these rats was measured by prelabeling cortical slices with myo-[2-3H]inositol and was expressed as fractional release (FR) of inositol (mono-, bis-, and tris-) phosphates. Acute increase in sodium intake produced 93 +/- 8% increase over control in urinary DA excretion. These changes were accompanied by significant increases (30 +/- 8%) in basal FR of inositol phosphates and 243 +/- 40 and 76 +/- 14% increases in urinary sodium and water excretion, respectively. The elevated basal PLC activity in rats with high sodium intake was significantly reduced in the presence of Sch 23390, a selective DA-1 receptor antagonist. Exogenously added DA (3 mM) also produced significant increases in PLC activity, although the magnitudes of increases were different in rats with high (37 +/- 8%) and normal (66 +/- 9%) sodium intake. However, Sch 23390 alone or carbidopa pretreatment did not affect the basal PLC activity in rats maintained on normal sodium intake.(ABSTRACT TRUNCATED AT 250 WORDS)
Participation of protein kinase C in desensitization to bradykinin and to carbachol in MDCK cells
Authors: Luo H. , Lindeman R. P. , Chase H. S.
Published: December 2017

To explore the possibility that protein kinase C (PKC) participates in desensitization to Ca(2+)-mobilizing hormones in MDCK cells, we measured intracellular free Ca2+ concentration ([Ca2+]i) using fura-2 and video microscopy. We first examined the response of MDCK cells grown on plastic dishes. Exposure of cells to bradykinin (BK) or to carbachol, followed by reexposure after washing off the hormone, revealed two features of hormone desensitization. First, the initial hormone-induced peak response of [Ca2+]i was transitory; [Ca2+]i returned to control levels despite continued presence of hormone. Second, cells remained refractory to hormone rechallenge for 5 min after washing off hormone; [Ca2+]i response on re-exposure was reduced 70% compared with initial hormone-stimulated peak. Subsequent experiments demonstrated involvement of PKC in both desensitization processes. Pretreatment with the phorbol ester, phorbol 12-myristate 13-acetate, significantly blunted initial response to BK and to carbachol by 70 and 86%, respectively. When hormone-stimulated C kinase activity was enhanced with the diglyceride lipase inhibitor, RG 80267, BK- and carbachol-induced increases in [Ca2+]i were blunted 50%. Pretreatment with sphingosine, an inhibitor of PKC, resulted in an amplification of initial hormone-stimulated increase in [Ca2+]i and restored the response to rechallenge. To examine the possible interaction between BK and carbachol,both of which use PKC to induce desensitization, we measured [Ca2+]i in cells grown as monolayers on permeable, collagen-coated supports. Both carbachol and BK induced desensitization to the other hormone (heterologous desensitization)provided that the two hormones were applied to the same side of the polarized monolayer (apical).(ABSTRACT TRUNCATED AT 250 WORDS)
Roles of cephalic Na+ concentration and urodilatin in control of renal Na+ excretion
Authors: Emmeluth C. , Drummer C. , Gerzer R. , Bie P.
Published: December 2017

Effects on renal function of an increase in the concentration of sodium in the blood supplying the head were investigated in water-diuretic conscious dogs in which the sodium and water contents were controlled by separate servo-mechanisms. A selective 2% increase in the sodium concentration of the carotid blood was achieved by a split-infusion technique including infusions of hypertonic saline into both carotid arteries and water into a jugular vein at rates making the combined infusate isotonic. This procedure caused a 34-fold increase in renal sodium excretion concomitant with a fourfold increase in the rate of urinary excretion of urodilatin. A comparable isotonic volume expansion (isotonic saline infusion into carotid arteries and jugular vein) caused a significantly smaller (13-fold) increase in urinary rate of excretion of sodium (P less than 0.02) and no increase at all in the excretion of urodilatin. It is hypothesized that cephalic sodium concentration receptors regulate the rate of excretion of sodium via urodilatin even under the present slightly hypotonic conditions.
Metanephric transforming growth factor-alpha is required for renal organogenesis in vitro
Authors: Rogers S. A. , Ryan G. , Hammerman M. R.
Published: December 2017

The role of transforming growth factor-alpha (TGF-alpha) in metanephric development was examined. Metanephroi were removed from 13-day-old rat embryos and grown in organ culture for up to 6 days using serum-free chemically defined media. During this time the metanephroi increased in size and morphological complexity. Messenger RNA for TGF-alpha was present in the renal anlage. Immunoreactive TGF-alpha was produced by metanephroi in vitro and released into culture media. TGF-alpha of metanephric origin coeluted with recombinant human 125I-TGF-alpha after high-performance liquid chromatography of media. In contrast, epidermal growth factor was not detectable. Levels of TGF-alpha were relatively constant during 4 days in culture and averaged approximately 10(-10) M. Growth of the metanephric anlage, arborization of the ureteric bud, and tubulogenesis within the metanephric blastema were inhibited by the addition of anti-TGF-alpha antibodies to organ cultures. These data demonstrate production of TGF-alpha by developing rat metanephroi in organ culture. The peptide is necessary for growth and development in vitro. Our findings suggest a necessary role for TGF-alpha of metanephric origin as a promoter of renal organogenesis in vivo.
Age-associated changes in ammoniagenesis in isolated rat renal tubule segments
Authors: Yamada H. , Sacktor B. , Kinsella J.
Published: December 2017

We investigated renal ammoniagenesis in isolated nephron segments from control and acidotic senescent and young adult rats. When young (6 mo) and senescent (24 mo) control groups were compared, there was no significant difference in glutamine-dependent ammonia production in any nephron tubule segments. However, ammonia production rates in glomeruli from old rats were significantly greater than the rate from young rats and were correlated with the serum creatinine and blood urea nitrogen (BUN) levels. After giving young and old rats an equivalent acid load (by gavage) of ammonia chloride solutions (6 mo, blood pH 7.34; 24 mo, blood pH 7.07), we measured a significant increase in ammoniagenesis on the S1 and S2 segments of proximal tubules and the distal convoluted tubules from old rats, and no increase in any segment from young rats. When we increased the acid load in young rats to an equivalent severity of acidosis (blood pH 7.07), we found significant increases in ammonia production in the S1, S2, S3, and distal convoluted tubule. With comparable blood pH values, ammoniagenesis in S1, S2, and S3 segments from young rats was about double the values measured in segments from senescent rats. The severity of the acidosis in the 24-mo-old rats was related to serum creatinine and BUN. Our findings show that ammoniagenesis in isolated segments from senescent rats is qualitatively similar to their younger counterparts but that this maximum capacity to generate ammonia is reduced. The change in nephron capacity to synthesize ammonia may be the result of age-associated physiological changes and/or the chronic renal failure exhibited in old rats.