bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of histidine on vasopressin action: role of decreased prostaglandin production
Authors: Carvounis C. P. , Schroeder E. T. , Cushley P. , Hueber P. , Patchin D.
Published: December 2017

Addition of histidine to the serosal bath of the toad bladder increases the hydrosmotic response of vasopressin in this tissue. Because this represents primarily the effect of the imidazole ring of histidine, which is a known inhibitor of the production of prostaglandins, we evaluated whether histidine increases the response to vasopressin through decreased prostaglandin production. Histidine increases the response to vasopressin much more than 10(-5) M naproxen, even though the latter was equipotent to histidine in reducing prostaglandin E2 (PGE2) production. Furthermore, histidine was additive to naproxen in increasing the hydrosmotic effect of vasopressin, without causing a further decrease in PGE2 production. These findings suggest that histidine has an effect over and above that due to inhibition of prostaglandin synthesis. Our results suggest that histidine enhances the permeability of the tissue beneath the luminal membrane, an effect not found with naproxen. We propose that histidine increases the hydrosmotic response to vasopressin through at least two distinct mechanisms: 1) it decreases prostaglandin synthesis and thus increases luminal permeability; 2) it decreases the resistance to water movement of the tissues beneath the luminal membrane.
Receptors and cGMP signalling mechanism for E. coli enterotoxin in opossum kidney
Authors: Forte L. R. , Krause W. J. , Freeman R. H.
Published: December 2017

Receptors for the heat-stable enterotoxin produced by Escherichia coli were found in the kidney and intestine of the North American opossum and in cultured renal cell lines. The enterotoxin markedly increased guanosine 3',5'-cyclic monophosphate (cGMP) production in slices of kidney cortex and medulla, in suspensions of intestinal mucosa, and in the opossum kidney (OK) and rat kangaroo kidney (PtK-2) cell lines. In contrast, atrial natriuretic factor elicited much smaller increases in cGMP levels of kidney, intestine, or cultured kidney cell lines. The enterotoxin receptors in OK cells had a molecular mass of approximately 120 kDa when measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of receptors crosslinked with 125I-enterotoxin. The occurrence of receptors for the E. coli peptide in OK implies that these receptors may be involved in the regulation of renal tubular function in the opossum. E. coli enterotoxin caused a much larger increase in urine cGMP excretion (10- to 50-fold over control) than did atrial natriuretic factor when these peptides were injected intravenously into opossums. However, atrial natriuretic factor elicited a marked diuresis, natriuresis, and increased urinary excretion of calcium, phosphate, potassium, and magnesium. In contrast, the enterotoxin did not acutely influence OK fluid and electrolyte excretion. Thus the substantial increase in cGMP synthesis produced by the bacterial peptide in OK cortex and medulla in vitro and the increased renal excretion of cGMP in vivo were not associated with changes in electrolyte or water excretion. Whether cGMP represents a second messenger molecule in the kidney is an interesting question that was raised but not answered in this series of experiments.
Ammonia loss from rat proximal tubule in vivo: effects of luminal pH and flow rate
Authors: Simon E. E. , Fry B. , Hering-Smith K. , Hamm L. L.
Published: December 2017

The roles of luminal pH and flow rate in determining ammonia loss from proximal tubules perfused with solutions containing 10 mM NH4Cl were examined using in vivo microperfusion. Perfusate bicarbonate concentration was varied between 5, 25, and 40 mM in tubules perfused at 50 nl/min. As expected, ammonia loss from the 25 or 40 mM bicarbonate-containing perfusates was greater than from that containing 5 mM bicarbonate. Furthermore, there was a correlation between ammonia loss and the log mean luminal bicarbonate concentration (r = 0.39, P less than 0.01). From the collected fluid ammonia and bicarbonate concentrations, the transtubular gradients for NH+4 and NH3 were estimated, allowing a calculation of the apparent permeability coefficients for NH3 (PNH3) and NH+4 (PNH+4). The calculated PNH3 of 2.2 +/- 0.5 X 10(-2) cm/s was similar to previous estimates in the rabbit; the calculated PNH+4 of 5.5 +/- 0.8 X 10(-4) cm/s was approximately 10 times that previously found in the rabbit proximal straight tubule in vitro. Next, flow rate was varied between 25 and 50 nl/min using the 5 mM bicarbonate perfusate. Ammonia loss was significantly higher from the latter. Thus these studies suggest that NH+4 loss from the proximal tubule may be an important determinant of ammonia movement along this segment. Ammonia loss is flow-rate dependent, similar to ammonia entry in previous studies.
Substrate specificity to maintain cellular ATP along the mouse nephron
Authors: Uchida S. , Endou H.
Published: December 2017

To evaluate substrate utilization along the mouse nephron, cellular ATP content was measured under various conditions by the luciferin-luciferase technique. Individual micro-dissected nephron segments were incubated in a modified Hanks' solution (pH 7.4) with or without each of the following substrates: D-glucose, DL-lactate, beta-hydroxybutyrate (HBA), and L-glutamine. ATP production from glucose was minimal in the early proximal tubule (S1), but was substantial in the late proximal tubule (S3). Glutamine and lactate were the preferred substrates in proximal tubules. In contrast, ATP production from glutamine was less than that from the other substrates in distal nephron segments, including medullary and cortical thick ascending limbs of Henle's loop (MTAL and CTAL), distal tubule including the connecting tubule (DT), and cortical and medullary collecting tubules (CCT and MCT). Lactate and HBA were preferred substrates for ATP maintenance in CTAL, MTAL, and DT. Glucose was the best substrate in CCT. In addition, the specific contribution of anaerobic metabolism to maintaining cellular ATP was low in MTAL and CTAL. On the other hand, the glycolytic capacity of CCT and MCT was high. The above results demonstrate the substrate requirements for maintaining cellular ATP content within specific nephron segments.
Mechanism of phosphaturia elicited by administration of phosphonoformate in vivo
Authors: VanScoy M. , Loghman-Adham M. , Onsgard M. , Szczepanska-Konkel M. , Homma S. , Knox F. G. , Dousa T. P.
Published: December 2017

We examined whether phosphonoformate (PFA) can cause phosphaturia through its direct action on brush-border membrane (BBM) in vivo. Infusion of PFA or of parathyroid hormone (PTH) to thyroparathyroidectomized rats caused a marked increase in fractional excretion of phosphate without changes in excretion of Na+ or of GFR. The PFA-induced phosphaturia was not accompanied by an increase in urinary adenosine-3',5'-cyclic monophosphate (cAMP); moreover, PFA added in vitro did not influence the PTH-sensitive adenylate cyclase and cAMP-phosphodiesterase in proximal convoluted tubules. In BBM vesicles (BBMV) from rats with PFA-elicited phosphaturia, neither the rate of Na+-Pi symport nor Na+-dependent binding of [14C]PFA on BBMV was changed, whereas in BBMV from PTH-infused rats the Vmax of Na+-Pi symport decreased. PFA is almost completely ultrafiltrable; no metabolic transformation of PFA was detected after [14C]PFA exposure to rat renal cortical slices, homogenate, or to blood. We conclude that PFA causes phosphaturia by direct inhibition of Na+-Pi symport across BBM in proximal tubules, acting from the luminal side. Thus PFA (foscarnet) has a unique direct mechanism of phosphaturic effect, via its action on Pi reabsorption in proximal tubules in vivo.
Alpha 1-adrenoceptor mediation of urinary bicarbonate excretion
Authors: Osborn J. L. , Harland R. W.
Published: December 2017

The adrenoceptor mechanism that mediates the reduction of urinary sodium (UNa V) and bicarbonate (UHCO3 V) excretion during 1.0-Hz renal nerve stimulation (RNS) was evaluated in 21 anesthetized dogs. In each animal RNS decreased UNa V and UHCO3 V without changing mean arterial pressure, renal blood flow, or glomerular filtration rate. After these initial responses, dogs were administered phentolamine [2.0 micrograms.kg-1.min-1 intrarenal artery (ira); n = 7], prazosin (0.7 micrograms.kg-1.min-1 ira; n = 5), (+/- )-propranolol (2.0 micrograms.kg-1.min-1 ira; n = 4), or atenolol (50 mg/kg iv; n = 5) and the renal responses to RNS were again tested. Both the antinatriuretic response and reduction of UHCO3 V during RNS were abolished by alpha 1/alpha 2-adrenoceptor blockade with phentolamine and by alpha 1-adrenoceptor antagonism with prazosin. beta-Adrenoceptor blockade with (+/- )-propranolol (beta 1/beta 2) or atenolol (beta 1) did not alter either the decrease in UNa V or UHCO3 V after RNS. These results provide further evidence that, during low-frequency RNS, reduction of UNa V is mediated in part by carbonic anhydrase-dependent bicarbonate reabsorption. These antinatriuretic responses and decreases in UHCO3 V during 1.0-Hz RNS are mediated by adrenergic neurotransmitter stimulation of alpha 1-adrenoceptors.
Succinate alters respiration, membrane potential, and intracellular K+ in proximal tubule
Authors: Gullans S. R. , Kone B. C. , Avison M. J. , Giebisch G.
Published: December 2017

Succinate, a dicarboxylic acid, is an intermediate in the Krebs cycle that is transported and metabolized by the renal proximal tubule. It is also known to increase proximal tubule transport of phosphate and glucose but not fluid by unknown mechanisms. In the present study, succinate increased proximal tubule respiration in a dose-dependent manner, and a kinetic evaluation indicated that two separate processes were activated. A lower-affinity (Km = 0.9 mM), higher-capacity stimulation (Vmax increase of 49%) was attributed to a decrease in the mitochondrial coupling efficiency. A higher-affinity process (Km = 0.012 mM) was related to an apparent increase in ATP synthesis. The apparent increase in ATP synthesis was not associated with a change in Na+-K+-ATPase activity, however, but rather indicated a 49% increase in ion transport-independent ATP utilization. Basolateral membrane potential hyperpolarized by -7 mV in the presence of succinate, and this was related to an increase in the K+ transference number. Finally, 1 and 5 mM succinate promoted a net cellular uptake of K+, leading to an 11% increase in intracellular K+, which was not the result of an increase in Na+-K+-ATPase activity. Thus the cellular entry and metabolism of succinate promotes multiple changes in ion transport without altering Na+-K+-ATPase activity.
Crystal adsorption and growth slowing by nephrocalcin, albumin, and Tamm-Horsfall protein
Authors: Worcester E. M. , Nakagawa Y. , Wabner C. L. , Kumar S. , Coe F. L.
Published: December 2017

Urine inhibition of calcium oxalate monohydrate (COM) crystal growth (CG) seems due to a glycoprotein that contains gamma-carboxyglutamic acid and has been named nephrocalcin (NC); however, Tamm-Horsfall protein (THP) and albumin resemble NC and make its measurement and role uncertain. NC in urine is aggregated to molecular mass 64 kDa and higher, similar to albumin (64 kDa) and THP (87 kDa). Albumin and THP are calcium binding, albumin adsorbs to COM crystals, and THP has been described as an inhibitor of COM growth. Antisera to NC have cross-reacted with THP even though the NC was isolated from cultured renal cells. Here we have compared highly purified NC, THP, and albumin adsorption with COM crystals and CG inhibition; also we compared their patterns of cross-reactivities with a new antiserum against NC and a monoclonal antibody to THP. NC adsorbs to COM crystals, THP does not. Albumin and THP do not inhibit CG. Cross-reactivity of albumin and THP to the antiserum is slight by direct enzyme-linked immunosorbent assay and nonexistent by competitive ELISA; reaction of NC to the anti-THP monoclonal antibody is absent.
Insulin increases loop segment chloride reabsorption in the euglycemic rat
Authors: Kirchner K. A.
Published: December 2017

To elucidate the nephron segment(s) responsible for insulin's antinatriuretic effect in the mammalian kidney, late proximal, early distal, and late distal tubule micropuncture was performed during base-line and intravenous insulin infusion in glucose-clamped, volume-expanded rats. During insulin infusion, urinary sodium chloride excretion fell. Blood glucose, arterial pressure, whole kidney and single-nephron inulin clearance, fluid, and chloride delivery out of the proximal convoluted tubule remained unchanged. Early distal chloride delivery decreased (P less than 0.001) during insulin administration. Insulin infusion increased calculated loop chloride reabsorption compared with both base-line values and values for time-control rats. Distal convoluted tubule chloride reabsorption was unchanged during insulin administration. We conclude that 1) euglycemic insulin administration reduces urinary chloride excretion in volume-expanded rats by tubular mechanisms; 2) in superficial nephrons, insulin has no effect on proximal tubule fluid or chloride reabsorption but markedly stimulates chloride reabsorption in the loop; and 3) chloride reabsorption in the distal convoluted tubule was unaltered by insulin administration.
Effect of spironolactone on glucocorticoid-induced colonic cation transport
Authors: Bastl C. P.
Published: December 2017

Glucocorticoids stimulate colonic sodium absorption. Whether this stimulation is mediated by crossover binding to the aldosterone receptor has not been established. The effect of the aldosterone receptor antagonist, spironolactone, on dexamethasone-mediated transport was investigated in proximal and distal colon of adrenalectomized rats. In adrenal-intact control rats spironolactone minimally affected sodium absorption but markedly decreased the transmural potential difference (PD). In adrenalectomized, aldosterone-replaced rats spironolactone clearly decreased sodium absorption and transmural PD in both proximal and distal colon. In adrenalectomized rats treated with 5 micrograms/100 g body wt of dexamethasone twice daily spironolactone did not decrease sodium absorption or transmural PD. However, when adrenalectomized rats were treated with 25 micrograms/100 g body wt of dexamethasone, a dose estimated from plasma levels to occupy 80% of colonic aldosterone receptors, spironolactone inhibited 20% of proximal colon and 47% of distal colon sodium absorption. Thus lower doses of glucocorticoid which predominantly occupy glucocorticoid receptors stimulate sodium absorption by a spironolactone independent mechanism. When the levels of glucocorticoid are high enough to also occupy the majority of aldosterone receptors, sodium absorption is stimulated by an aldosterone receptor mediated mechanism.
Increased single-nephron protein excretion after renal ablation in nephrotic rats
Authors: Meyer T. W. , Rennke H. G.
Published: December 2017

The effects of reducing nephron number in rats with established nephrosis were investigated. Rats received an injection of adriamycin and were divided into three groups with similar values for proteinuria after 4 wk. Group 1 rats were then subjected to sham operation. Group 2 rats were subjected to four-fifths renal ablation, and group 3 rats were subjected to four-fifths renal ablation and then maintained on enalapril. Micropuncture and morphological studies were performed 3 wk later. During this 3-wk period, proteinuria increased slightly in each group. Increased proteinuria in groups 2 and 3 reflected a dramatic increase in remnant nephron proteinuria after renal ablation in nephrotic rats. Increased remnant nephron proteinuria in groups 2 and 3 was associated with increased single-nephron glomerular filtration rate (group 1, 30 +/- 2 nl/min; group 2, 54 +/- 3 nl/min; group 3, 41 +/- 4 nl/min) and increased glomerular volume (group 1, 0.93 +/- 0.05 x 10(6) micron 3; group 2, 1.30 +/- 0.09 x 10(6) micron 3; group 3, 1.27 +/- 0.05 x 10(6) micron 3). The increase in single-nephron glomerular filtration rate after renal ablation in both group 2 and 3 rats was attributable to an increase in glomerular plasma flow (group 1, 119 +/- 14 nl/min; group 2, 217 +/- 18 nl/min; group 3, 183 +/- 13 nl/min) without a significant increase in glomerular transcapillary hydraulic pressure (group 1, 45 +/- 1 mmHg; group 2, 48 +/- 3 mmHg; group 3, 44 +/- 2 mmHg). Group 2 exhibited an increase in systemic blood pressure that was prevented by enalapril treatment in group 3. These studies show that an increase in remnant nephron proteinuria accompanies glomerular hypertrophy and hyperfiltration when nephron number is reduced in nephrotic rats. This increase in remnant nephron proteinuria is not attributable to elevation of systemic or glomerular capillary pressure.
LLC-PK1 cells express Na+-lactate cotransport in apical membranes after confluency
Authors: Poustis-Delpont C. , Mengual R. , Sudaka P.
Published: December 2017

L-[3H]lactate uptake was characterized in LLC-PK1 cell apical membrane vesicles obtained by intensive culture on microcarrier beads. The apical membrane preparation technique involved MgCl2 precipitation. Na+-dependent L-[3H]lactate uptake was present only after confluency; its appearance paralleled the subcellular localization of aminopeptidase in apical membranes. L-[3H]lactate uptake was Na+-dependent and electrogenic. Only the Na+-dependent component of L-[3H]lactate uptake was saturable with one family of independent carriers. The apparent affinity constant was 1.1 +/- 0.25 mM and the apparent maximal velocity was 29 +/- 3 nmol.mg-1.min-1. The Na+-lactate cotransport stoichiometry was 2 Na+ for 1 lactate. The specificity of the L-lactate transport system was compatible with that of the monocarboxylic acid pathway described previously in brush-border membranes of kidney cortex and discrete from the tricarboxylic acid carrier, the D-glucose transporter, and the general pathway for anions. The LLC-PK1 cell line appears to be a useful tool for study of the regulation of L-lactate uptake and biosynthesis of the renal monocarboxylic acid transporter.
Renal inner medullary choline dehydrogenase activity: characterization and modulation
Authors: Grossman E. B. , Hebert S. C.
Published: December 2017

Betaine belongs to the trimethylamine class of osmolytes (osmotically active substances believed to play an important role in cell volume homeostasis) and has recently been identified in the inner medulla of the mammalian kidney. Trimethylamines accumulate in the renal inner medulla during hypertonic stress, and betaine content in the inner medulla has been shown recently to increase during hypernatremia, yet the mechanisms governing the modulation of trimethylamine content and, in particular, of betaine content are not well understood. In this study, we demonstrate the presence of choline dehydrogenase activity in the renal inner medullas of three separate rat strains. Choline dehydrogenase is the enzyme that catalyzes the first of two successive oxidation steps in the biosynthetic conversion of choline to betaine. The presence of choline dehydrogenase activity in the inner medulla suggests that betaine accumulation in the inner medulla may result, at least in part, through in situ synthesis. The Km and Vmax of the reaction in the inner medullas of Long-Evans rats are 4.7 +/- 0.5 mM and 36.9 +/- 5.0 nmol.mg protein-1.min-1, respectively. These values are similar to the characteristics of choline dehydrogenase in mammalian liver. During hypernatremia, when betaine content of the inner medulla has been shown to increase 1.5-fold, choline dehydrogenase activity remains unchanged (or slightly increased), whereas enzyme activity in the cortex increases approximately 50%. Possible mechanisms of inner medullary betaine accumulation are discussed.
Renal microvascular effects of vasopressin and vasopressin antagonists
Authors: Edwards R. M. , Trizna W. , Kinter L. B.
Published: December 2017

The effects of vasopressin (AVP) and vasopressin antagonists on lumen diameters of cortical afferent and efferent arterioles isolated from rabbit kidneys were examined. Over a concentration range of 10(-14) to 10(-7) M, AVP had no effect on lumen diameters of afferent arterioles, although the arterioles were responsive to norepinephrine. Similarly, addition of 10(-8) M AVP to the lumen of afferent arterioles or to the bath of arterioles pretreated with indomethacin had no effect. In contrast, AVP caused a concentration-dependent reduction of lumen diameters of efferent arterioles. AVP was approximately 100-fold more potent than norepinephrine in producing contraction of efferent arterioles. The V1-selective antagonist, [d(CH2)5Tyr(Me)]AVP, and the V1/V2-antagonist, d(CH2)5D-Tyr(Et) desGlyVAVP, inhibited the vasoconstriction produced by AVP in a concentration-dependent but noncompetitive manner. The V2-selective antagonist, [d(CH2)5D-Ile]VAVP, had no significant effect on AVP-induced vasoconstriction. We conclude that, under the in vitro conditions used, AVP selectively contracts efferent arterioles. The results provide direct evidence for a postglomerular vascular effect of AVP in the renal cortex. This activity, together with its previously described effects on the glomerulus, suggests that AVP may produce changes in glomerular function and/or peritubular forces that are involved in tubular reabsorption.
Depolarization-induced alkalinization in proximal tubules. I. Characteristics and dependence on Na+
Authors: Siebens A. W. , Boron W. F.
Published: December 2017

We used intracellular pH-sensitive and voltage microelectrodes to examine the effects of depolarization on intracellular pH (pHi) in isolated perfused proximal tubules from the tiger salamander Ambystoma tigrinum. Tubules were depolarized by raising [K+] in the bath (b) or lumen (l), or by adding Ba2+ (1 mM) to the bath or lumen, always in nominally HCO3-free solutions. Increasing [K+]b from 2.5 to 50 mM caused the basolateral membrane to depolarize by an average of 45 mV, and pHi to increase by 0.23 over 3 min. Similar alkalinization was observed when basolateral Ba2+ (1 mM) was used to depolarize the cell at constant extracellular [K+], suggesting that the alkalinization observed during exposure to elevated [K+]b results from depolarization rather than an increase in [K+]b. The initial rate of depolarization-induced alkalinization (DIA) was proportional to the magnitude of the depolarization, regardless of whether tubules were depolarized by elevated [K+]b, elevated [K+]l, or by basolateral Ba2+. An exception was the initial rate of the alkalinization caused by 1 mM luminal Ba2+, which was more than 10-fold greater than that predicted from the depolarization. The voltage and pHi responses to basolateral Ba2+ were smaller in some tubules than others, as were the responses to elevated [K+]l. Tubules with small responses to 1 mM [Ba2+]b had large responses to 50 mM [K+]l, whereas tubules with large responses to 1 mM [Ba2+]b had small responses to 50 mM [K+]l. This variability can be accounted for by differences in the luminal K+ conductance. The DIA was partially inhibited by removal of Na+ from only the lumen or only the bath, but completely inhibited by bilateral Na+ removal. We conclude that the depolarization-induced alkalinization results from additive effects of Na+-dependent processes at both the luminal and basolateral membranes.
Immunocytochemical localization of Na+ channels in rat kidney medulla
Authors: Brown D. , Sorscher E. J. , Ausiello D. A. , Benos D. J.
Published: December 2017

Amiloride-sensitive Na+ channels were localized in semithin frozen sections of rat renal medullary collecting ducts, using polyclonal antibodies directed against purified bovine kidney Na+ channel protein. The apical plasma membrane of collecting duct principal cells was heavily stained by indirect immunofluorescence, whereas intercalated cells were negative. Basolateral plasma membranes of both cell types were unstained, as were subapical vesicles in the cytoplasm of these cells. In the thick ascending limb of Henle, some scattered granular fluorescence was seen in the cytoplasm and close to the apical pole of epithelial cells, suggesting the presence of antigenic sites associated with some membrane domains in these cells. No staining was detected in thin limbs of Henle, or in proximal tubules in the outer medulla. These results show that amiloride-sensitive sodium channels are located predominantly on the apical plasma membrane of medullary collecting duct principal cells, the cells that are involved in Na+ homeostasis in this region of the kidney.
Effect of dietary K+ and 75% nephrectomy on the morphology of principal cells in CCDs
Authors: Zalups R. K.
Published: December 2017

This investigation was carried out to determine the effect of potassium on the amplification of the basolateral membrane of principal cells in cortical collecting ducts (CCDs), which normally occurs after a large reduction of renal mass. Rats that underwent a 75% reduction of renal mass and were fed a diet deficient in potassium for 10 days after surgery had a lower concentration of potassium in the blood and excreted less potassium in the urine than either corresponding sham-operated (SO) rats or 75% nephrectomized (NPX) rats fed a normal-potassium diet. In the NPX rats fed the diet deficient in potassium, there was a marked inhibitory effect on the amplification of the basolateral membrane, which normally occurs in principal cells of CCDs after renal mass has been greatly reduced. In fact the surface density of the basolateral membrane, as well as the size of the principal cells in the animals fed the diet deficient in potassium were the same as those of principal cells in the SO rats fed the normal-potassium diet. Thus the present data show that the increase in cell size and the amplification of the basolateral membrane that occurs in principal cells after a 75% reduction of renal mass is inhibited by factors associated with potassium depletion.
Epithelial polarity following ischemia: a requirement for normal cell function
Authors: Spiegel D. M. , Wilson P. D. , Molitoris B. A.
Published: December 2017

Ischemic injury results in proximal tubule (PT) dysfunction and loss of surface membrane (SM) polarity. Since epithelial vectorial transport requires SM polarity, we set out to determine if correction of renal cortical PT dysfunction following ischemia was dependent on the reestablishment of SM polarity. Acute renal failure was induced using a bilateral 50-min pedicle clamp. Serum creatinine and fractional sodium excretion were maximal on day 1, remained elevated on day 3, and returned toward base line by day 8. PT cellular ultrastructure was normal by day 3. Despite rapid morphological recovery, ischemia resulted in a prolonged defect in glucose reabsorption. The delayed recovery of normal glucose handling closely paralleled the slow normalization of apical membrane lipid polarity. Na+-K+-ATPase polarity was also lost secondary to ischemia as demonstrated cytochemically and biochemically by the redistribution of Na+-K+-ATPase to the apical membrane. The time required to reestablish normal Na+-K+-ATPase polarity (8 days) paralleled the recovery of normal PT Na+ reabsorption (8 days), as assessed by fractional lithium clearances. This finding supports the hypothesis that apical Na+-K+-ATPase is in part responsible for reduced Na+ reabsorption following ischemic injury. In summary, these data suggest that functional recovery of PT glucose and Na+ reabsorption following a reversible ischemic insult requires not only morphological recovery, but also the reestablishment of surface membrane lipid and protein polarity.
Altered sodium regulation of renal angiotensinogen mRNA in the spontaneously hypertensive rat
Authors: Pratt R. E. , Zou W. M. , Naftilan A. J. , Ingelfinger J. R. , Dzau V. J.
Published: December 2017

The expression of the intrarenal renin angiotensin mRNAs in Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) were investigated in rats fed a low (0.02%)- or normal (1.6%)-sodium diet for 5 days. Total RNA was isolated from the kidneys and analyzed by both Northern and slot blots. The results indicated that the kidneys of all four groups of rats containing readibly detectable levels of renin and angiotensinogen mRNAs. The kidneys of the WKY contained higher levels of angiotensinogen mRNA under normal-salt diet compared with the SHR (P less than 0.01). Mild sodium depletion stimulated angiotensinogen mRNA in WKY kidneys by almost 50% compared with normal salt diet (P less than 0.01). In contrast, there was no significant difference in the renal angiotensinogen mRNA levels in the kidneys of SHR fed a low- or normal-sodium diet. Renin mRNA concentrations were comparable in both strains. Mild sodium depletion failed to yield any detectable changes in renin mRNA levels in either strain. Thus the SHR exhibits an alteration in the sodium regulation of intrarenal angiotensinogen mRNA expression. These results may have implications in the renal physiology of these animals.
Interstitial dynamics in rats with early stage experimental cirrhosis of the liver
Authors: Sanz E. , Caramelo C. , Lopez-Novoa J. M.
Published: December 2017

Pathogenesis of edema in cirrhosis of the liver is still incompletely understood. The present study was designed to examine interstitial fluid dynamics in cirrhotic, non-ascitic rats, measuring interstitial fluid pressure by means of a subcutaneous plastic capsule in basal conditions during extracellular fluid volume expansion with Ringer solution and during albumin infusion. Urine flow and sodium excretion and plasma and interstitial fluid volumes were simultaneously measured. Cirrhotic rats exhibited reduced urine flow and sodium excretion, both in basal conditions and in response to expansion maneuvers. Plasma and interstitial fluid volumes were higher in cirrhotic than in control animals. Remarkable alterations were present in capsular pressures in cirrhotic rats. In the control rats, basal capsular pressure values were negative, and they increased after Ringer infusion and markedly decreased with albumin infusion. In contrast, in cirrhotic rats, basal capsular pressures were in the positive range and they remained nearly constant during ringer infusion and albumin administration. These results suggest that in cirrhotic rats there are significant alterations in systemic interstitial dynamics, even before ascites formation. Altered systemic capillary dynamics may therefore be important early changes that precede and thus contribute to the formation of edema in cirrhosis.
Variability of cellular responsiveness to ADH stimulation in toad urinary bladder
Authors: Kachadorian W. A. , Muller J. , DiScala V. A.
Published: December 2017

The hydrosmotic response of toad bladder to antidiuretic hormone (ADH) is quantitatively linked to the induced fusion of aggrephores with, and the appearance of aggregates of tightly packed intramembrane particles in the luminal membrane of granular cells. We used these morphological indexes of hormonally induced cell activation 1) to assess the variability of individual cell responsiveness to a maximally stimulating concentration of ADH and 2) to compare cell response patterns in paired tissues where the extent of whole tissue stimulation, as evidenced by transtissue water flow, was either maximal or submaximal. The results indicate that individual cell responsiveness within the same tissue to standardized maximal ADH treatment varies between two- and sevenfold, depending on the morphological endpoint measured. Furthermore, based on skewness in endpoint distribution, this variability appears to reflect inherent heterogeneity of granular cell reactivity to hormone. In relation to proportional tissue responses elicited by different stimulating concentrations of ADH, our observations of luminal membrane aggregate incidence suggest that the responding cells, whatever their sensitivity, participate in a graded, rather than "all-or-none," "on-off" manner.
Independence of urea and water transport in rat inner medullary collecting duct
Authors: Knepper M. A. , Sands J. M. , Chou C. L.
Published: December 2017

Several published models of the renal concentration mechanism have assumed a reflection coefficient for urea in the inner medullary collecting duct (IMCD) that is less than unity, implying direct coupling between water and urea transport. In the present study, we used isolated perfused terminal IMCD segments and mathematical modeling of IMCD transport to determine the validity of this assumption. Mathematical simulations of IMCD transport, using recently published data on urea and water permeability, revealed that the method previously used to measure the reflection coefficient for urea underestimates the true value. The modeling results allowed us to design two new experimental protocols to determine the reflection coefficient for urea. In the first protocol, we measured the ability of a transepithelial urea gradient to induce a water flux, correcting for the dissipation of the urea gradient by rapid passive urea permeation. In the second protocol, we directly measured the solvent drag of urea resulting from an osmotically induced water flux. Both protocols yielded values for the urea reflection coefficient that were not significantly different from unity (0.92 +/- 0.04 and 1.07 +/- 0.05, respectively). Thus we find no evidence for direct coupling between urea and water transport in the rat terminal IMCD.
The role of free fatty acids in hypoxia-induced injury to renal proximal tubule cells
Authors: Humes H. D. , Nguyen V. D. , Cieslinski D. A. , Messana J. M.
Published: December 2017

Phospholipase activation with resulting phospholipid breakdown and lipid byproduct accumulation may play a critical role in hypoxic cell injury. To explore this role, mildly hypoxic rabbit renal proximal tubules (PT) in suspension were treated in vitro with exogenous phospholipase A2 (PLA2). This treatment produced severe tubule cell injury measured by alterations in tubule cation homeostasis, respiratory rates, and adenosine nucleotide metabolism. This injury was associated with loss of the major membrane phospholipids, phosphatidylcholine (PC) and phosphatidylethanolamine (PE), with accumulation of lipid byproducts, lysophosphatidylcholine (LPC), lysophosphatidylethanolamine (LPE), and free fatty acids (FFA). Addition of fatty acid-free bovine serum albumin (BSA) to PTs reduced markedly FFA levels and improved significantly derangements in metabolic parameters of hypoxic PTs treated with exogenous PLA2, suggesting that FFA accumulation was a critical factor in this injury process. Effects of increasing durations of hypoxia (30, 45, and 60 min) with or without reoxygenation recovery demonstrated increased FFA levels, especially polyunsaturated FFA, which correlated better with the degree of hypoxic injury than alterations in membrane phospholipid and lysophospholipid levels. PTs undergoing hypoxia and reoxygenation recovery exposed to BSA were not protected. Although 60 min of hypoxia with 60 min reoxygenation produced accumulation of FFA to levels nearly identical to those seen in hypoxic PTs treated with exogenous PLA2 and BSA, with a similar distribution of various FFA species, hypoxia/reoxygenation produced a more severe degree of cell injury than that observed with hypoxia plus exogenous PLA and BSA.(ABSTRACT TRUNCATED AT 250 WORDS)
Isolated principal and intercalated cells: hormone responsiveness and Na+-K+-ATPase activity
Authors: Fejes-Toth G. , Naray-Fejes-Toth A.
Published: December 2017

To explore biochemical and functional differences between principal (PC) and intercalated cells (ICC), we have developed a method for separating them from rabbit kidney. Fragments of cortical collecting ducts were isolated by immunodissection, and single cells obtained from these clusters were stained with fluorochrome-conjugated, cell-specific markers. PC and ICC were then separated by fluorescence-activated cell sorting. Identity of the sorted cells was confirmed by staining with other cell-specific monoclonal antibodies (MCABs) or peanut lectin. Purity was greater than 99% for ICC and greater than 96% for PC. Arginine vasopressin (AVP) increased adenosine 3',5'-cyclic monophosphate (cAMP) accumulation in both cell types, but maximal stimulation was significantly greater with PC (approximately 20-fold) than with ICC (2.7-fold). Half-maximal stimulation was seen at approximately 2 x 10(-10) M AVP with both cell types. Isoproterenol increased cAMP levels only with ICC (from 1.23 +/- 0.16 to 12.06 +/- 1.25 fmol/cell; P less than 0.001). The number of ouabain binding sites and the activity of Na+-K+-ATPase was significantly higher in sorted PC than ICC (2.2 X 10(6) vs. 9.6 X 10(5) binding sites; 19.2 vs. 9.6 fmol.min-1.cell-1 ATP hydrolyzed in PC vs. ICC, respectively). These results demonstrate the feasibility of isolating homogeneous populations of PC and ICC, which is useful for further studies of their biochemical and functional characterization.
Micropuncture localization of the natriuretic effect of interleukin 1
Authors: Kohan D. E. , Merli C. A. , Simon E. E.
Published: December 2017

Interleukin 1 (IL-1) has been demonstrated to elicit an increase in renal sodium excretion. This effect occurs in the absence of any increase in the filtered load of sodium, raising the possibility of an IL-1-mediated decrease in tubule sodium reabsorption. To localize the nephron segment(s) responsible for the natriuretic effect of IL-1, we performed micropuncture experiments on rats. Intravenous IL-1 administration caused a marked increase in sodium excretion that was not accompanied by changes in glomerular filtration rate or systemic blood pressure. Single-nephron glomerular filtration rate and fractional and absolute delivery of sodium to the late proximal and mid-distal tubule were not affected by IL-1. Fractional delivery of sodium to the early and late papillary collecting duct, however, was significantly enhanced by IL-1 administration. Sodium reabsorption was inhibited along the papillary collecting duct. These findings demonstrate that the natriuretic effect of IL-1 is due, at least in part, to inhibition of collecting duct sodium reabsorption.
Phorbol esters and arginine vasopressin in vascular smooth muscle cell activation
Authors: Caramelo C. , Okada K. , Tsai P. , Schrier R. W.
Published: December 2017

Vascular smooth muscle cell (VSMC) contraction is the result of several interacting mechanisms. In this study such interactions in rat aortic VSMC in culture were examined with a focus on the role of protein kinase C-mediated mechanisms. The change in shape of VSMC was used as a functional parameter representative of contraction. The protein kinase C agonist, phorbol myristate acetate (PMA), and arginine vasopressin (AVP) induced a dose-dependent, progressive change in VSMC shape. The effects of PMA differed from AVP in the delay time necessary to reach the plateau of the response and in the absence with PMA of a transient rise in cytosolic free Ca2+ [( Ca2+]i). The effect of PMA was potentiated by the Ca2+ ionophore, ionomycin, and involved a verapamil-inhibitable transmembrane Ca2+ transport system. Protein kinase C inhibition by either isoquinolin-sulfonyl-O-2-methylpiperazine or protein kinase C desensitization significantly reduced the cell shape change induced by either PMA or AVP. In the case of AVP, this inhibitory effect occurred without affecting the [Ca2+]i transient. Therefore, the [Ca2+]i transient appears to be independent of acute protein kinase C regulation, since it is apparently not affected by the absence of protein kinase C activity. Protein kinase C activation by PMA produced intracellular alkalinization that is blocked by the sodium transport antagonist, amiloride. Amiloride also blocked the cell shape change induced by PMA or AVP. The intracellular alkalinization, however, was not necessary for the cell shape change to occur. Specifically, with the use of VSMC preincubated with fetal calf serum, PMA did not induce cellular pH changes but still produced cell shape changes.
Characterization of glomerular thromboxane receptor sites in the rat
Authors: Wilkes B. M. , Solomon J. , Maita M. , Mento P. F.
Published: December 2017

The aim of this study was to identify and characterize thromboxane (Tx) receptor sites in renal glomeruli. Binding studies were performed on freshly isolated glomeruli using the stable TxA2 receptor antagonist, [3H]SQ 29548. Specific binding was saturable, reversible, and varied with glomerular protein. Scatchard plots revealed a single class of high-affinity receptor sites (Kd = 14.3 +/- 2.4 nM, Bmax = 361 +/- 22 fmol/mg; n = 5). Specific binding was inhibited by Tx agonists (U-46619 and U-44069) and antagonist (SQ 29548) and was highly specific for Tx, since prostaglandin (PG)E2 and PGF2 alpha were 1,000-fold less potent in inhibiting binding. In vivo, U-46619 (1.75 micrograms.kg-1.min-1) was without effect on mean arterial pressure, but reduced renal blood flow by 71% (P less than 0.01) and glomerular filtration rate by 67% (P less than 0.01) and increased filtration fraction by 24% (P less than 0.05). SQ 29548 (10 micrograms.kg-1.min-1) completely blocked the renal effects of U-46619. These studies demonstrate the presence of specific receptor sites for Tx on renal glomeruli that are linked to modulation of renal hemodynamics.
Differential effects of phorbol esters on PGE2 and bradykinin-induced elevation of [Cai2+] in MDCK cells
Authors: Aboolian A. , Vander Molen M. , Nord E. P.
Published: December 2017

Prostaglandin (PG) activation of the phosphoinositol transduction pathway in MDCK cells and modulation of this process by phorbol esters was studied by monitoring changes in cytosolic free Ca2+ concentration, [Cai2+], with the Ca2+-sensitive fluorescent probe, fura-2 and measurement of stimulation of inositol phosphates by anion-exchange chromatography. Cells challenged with PGE1 or PGE2 responded with a prompt and transient increase in [Cai2+] that was independent of extracellular Ca2+. The K0.5 for PGE2 for the process was 6.1 X 10(-7) M. PGE1 and PGE2 appeared to be recognized by a common receptor. PGF2 alpha was without effect. 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8) but not verapamil, a Ca2+ channel inhibitor, blocked the PGE2-evoked Ca2+ transient. Under identical conditions PGE2 increased inositol phosphate accumulation by 54 +/- 8% (inositol-1-monophosphate), 23 +/- 6% (inositol-1,4-bisphosphate), and 49 +/- 3% (total inositol trisphosphate), above control values. Brief (30-60 s) exposure of cells to phorbol-12,13-myristate (PMA) or phorbol-12,13-dibutyrate (PDB) completely blocked the PGE2-induced Ca2+ transient. The K0.5 for the process for PMA and PDB was 0.3 +/- 0.1 and 4.5 +/- 2.2 nM, respectively. Neither 4 alpha-nor 4 beta-phorbol, which lack the ability to activate protein kinase C, were effective in this regard. In contrast to complete blockade by 10(-8) PMA of the PGE2 (10(-5) M)-elicited Ca2+ transient, this concentration of PMA inhibited the Ca2+ transient evoked by 10(-9) M bradykinin (BK) by 50%. In fact 10(-4) M PMA only partially blocked the BK-elicited Ca2+ transient. In summary, in MDCK cells, the PG receptor is coupled both to the adenylate cyclase system and inositol phospholipid transduction pathway. The PG receptor appears to be regulated by protein kinase C. In addition to protein kinase C other factors regulate the BK receptor.
Peptide hormone effects on urinary acidification and acid-base balance: PTH, ADH, and glucagon
Authors: Paillard M. , Bichara M.
Published: December 2017

That the adaptation of the kidney to the acid-base status may be controlled by peptide hormones is considered. In the proximal tubule parathyroid hormone (PTH) inhibits reabsorption of both bicarbonate and phosphate. The former effect is compensated for by an increase in bicarbonate absorption in Henle's loop, and the latter effect serves to augment phosphate concentration in the distal tubular fluid, which stimulates proton secretion in collecting ducts, the net effect of PTH administration being an enhancement of urinary acidification. In the thick ascending limb, both antidiuretic hormone (ADH) and glucagon inhibit bicarbonate absorption. In distal and cortical collecting tubules ADH stimulates net bicarbonate absorption and glucagon net bicarbonate secretion, which results in stimulation and inhibition of final urine acidification, respectively. Acute acid loading stimulates endogenous PTH secretion, which, by enhancing urinary acidification, constitutes a homeostatic response of the parathyroid glands. The major effects of ADH on urinary acidification serve at least to counterbalance disturbing consequences on urinary ammonia excretion of physiological variations in the urinary flow rate. The physiological significance of the effects of glucagon is unclear at present. Thus other peptide hormones may add to PTH and corticosteroid hormones to modulate urinary acidification, which leads to the concept of a pluri-hormonal control of acid-base balance.
Antagonist to GH-releasing factor inhibits growth and renal Pi reabsorption in immature rats
Authors: Mulroney S. E. , Lumpkin M. D. , Haramati A.
Published: December 2017

Compared with adult rats, the immature rat has an enhanced tubular capacity for phosphate reabsorption, which presumably facilitates the growth process. Since the main driving force for growth is thought to be the pulsatile release of growth hormone, we examined the possibility that the adaptation in phosphate handling by the immature kidney is promoted by growth hormone (GH). To address this issue, we used a synthetic peptide antagonist to GH-releasing factor (GRF-AN) that we have shown blocks episodic GH secretion, and attenuates somatic growth. Immature male Wistar rats (4–5 wk of age) were catheterized with Silastic jugular cannulas and placed in metabolic cages. The rats were injected intravenously with either saline or GRF-AN (100 micrograms/kg) twice daily for 4 days. On the 4th day, they were prepared for renal clearance experiments to assess the maximum capacity for phosphate transport (TmPi). In animals treated with GRF-AN, there was an attenuated gain in body weight over 4 days of treatment (5 +/- 2 vs. 23 +/- 2% in saline controls, P less than 0.01). The suppressed growth was associated with a doubling of daily urinary phosphate excretion, and a reduction in the TmPi (3.3 +/- 0.1 vs. 4.6 +/- 0.3 mumol/ml in controls, P less than 0.01). A single injection of the antagonist to a separate group of immature rats did not alter TmPi. Thus injections of this new antagonist to GH-releasing factor over a 4-day period inhibit the pulsatile release of GH and significantly attenuate growth. The decline in growth of the immature rat was associated with a decrease in the renal capacity for phosphate reabsorption, down to levels seen in normal adult rats.(ABSTRACT TRUNCATED AT 250 WORDS)